@article{ReeveNicholsonAltafetal.2022,
  author    = {Reeve, Holly A. and Nicholson, Jake and Altaf, Farieha and Lonsdale, Thomas H. and Preissler, Janina and Lauterbach, Lars and Lenz, Oliver and Leimk{\"u}hler, Silke and Hollmann, Frank and Paul, Caroline E. and Vincent, Kylie A.},
  title     = {A hydrogen-driven biocatalytic approach to recycling synthetic analogues of NAD(P)H},
  series = {Chemical communications : ChemComm},
  volume    = {58},
  journal   = {Chemical communications : ChemComm},
  number    = {75},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  issn      = {1359-7345},
  doi       = {10.1039/d2cc02411j},
  pages     = {10540 -- 10543},
  year      = {2022},
  abstract  = {We demonstrate a recycling system for synthetic nicotinamide cofactor analogues using a soluble hydrogenase with turnover number of >1000 for reduction of the cofactor analogues by H-2. Coupling this system to an ene reductase, we show quantitative conversion of N-ethylmaleimide to N-ethylsuccinimide. The biocatalyst system retained >50\% activity after 7 h.},
  language  = {en}
}
@article{TeraoGarattiniRomaoetal.2020,
  author    = {Terao, Mineko and Garattini, Enrico and Rom{\~a}o, Maria Jo{\~a}o and Leimk{\"u}hler, Silke},
  title     = {Evolution, expression, and substrate specificities of aldehyde oxidase enzymes in eukaryotes},
  series = {The journal of biological chemistry},
  volume    = {295},
  journal   = {The journal of biological chemistry},
  number    = {16},
  publisher = {American Society for Biochemistry and Molecular Biology},
  address   = {Rockville},
  issn      = {0021-9258},
  doi       = {10.1074/jbc.REV119.007741},
  pages     = {5377 -- 5389},
  year      = {2020},
  abstract  = {Aldehyde oxidases (AOXs) are a small group of enzymes belonging to the larger family of molybdo-flavoenzymes, along with the well-characterized xanthine oxidoreductase. The two major types of reactions that are catalyzed by AOXs are the hydroxylation of heterocycles and the oxidation of aldehydes to their corresponding carboxylic acids. Different animal species have different complements of AOX genes. The two extremes are represented in humans and rodents; whereas the human genome contains a single active gene (AOX1), those of rodents, such as mice, are endowed with four genes (Aox1-4), clustering on the same chromosome, each encoding a functionally distinct AOX enzyme. It still remains enigmatic why some species have numerous AOX enzymes, whereas others harbor only one functional enzyme. At present, little is known about the physiological relevance of AOX enzymes in humans and their additional forms in other mammals. These enzymes are expressed in the liver and play an important role in the metabolisms of drugs and other xenobiotics. In this review, we discuss the expression, tissue-specific roles, and substrate specificities of the different mammalian AOX enzymes and highlight insights into their physiological roles.},
  language  = {en}
}
@misc{Leimkuehler2020,
  author    = {Leimk{\"u}hler, Silke},
  title     = {The biosynthesis of the molybdenum cofactors in Escherichia coli},
  series = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  number    = {6},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-51655},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-516559},
  pages     = {22},
  year      = {2020},
  abstract  = {The biosynthesis of the molybdenum cofactor (Moco) is highly conserved among all kingdoms of life. In all molybdoenzymes containing Moco, the molybdenum atom is coordinated to a dithiolene group present in the pterin-based 6-alkyl side chain of molybdopterin (MPT). In general, the biosynthesis of Moco can be divided into four steps in in bacteria: (i) the starting point is the formation of the cyclic pyranopterin monophosphate (cPMP) from 5 '-GTP, (ii) in the second step the two sulfur atoms are inserted into cPMP leading to the formation of MPT, (iii) in the third step the molybdenum atom is inserted into MPT to form Moco and (iv) in the fourth step bis-Mo-MPT is formed and an additional modification of Moco is possible with the attachment of a nucleotide (CMP or GMP) to the phosphate group of MPT, forming the dinucleotide variants of Moco. This review presents an update on the well-characterized Moco biosynthesis in the model organism Escherichia coli including novel discoveries from the recent years.},
  language  = {en}
}
@article{YanFrokjarEngelbrektetal.2021,
  author    = {Yan, Jiawei and Fr{\o}kj{\ae}r, Emil Egede and Engelbrekt, Christian and Leimk{\"u}hler, Silke and Ulstrup, Jens and Wollenberger, Ulla and Xiao, Xinxin and Zhang, Jingdong},
  title     = {Voltammetry and single-molecule in situ scanning tunnelling microscopy of the redox metalloenzyme human sulfite oxidase},
  series = {ChemElectroChem},
  volume    = {8},
  journal   = {ChemElectroChem},
  number    = {1},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {2196-0216},
  doi       = {10.1002/celc.202001258},
  pages     = {164 -- 171},
  year      = {2021},
  abstract  = {Human sulfite oxidase (hSO) is a homodimeric two-domain enzyme central in the biological sulfur cycle. A pyranopterin molybdenum cofactor (Moco) is the catalytic site and a heme b(5) group located in the N-terminal domain. The two domains are connected by a flexible linker region. Electrons produced at the Moco in sulfite oxidation, are relayed via heme b(5) to electron acceptors or an electrode surface. Inter-domain conformational changes between an open and a closed enzyme conformation, allowing "gated" electron transfer has been suggested. We first recorded cyclic voltammetry (CV) of hSO on single-crystal Au(111)-electrode surfaces modified by self-assembled monolayers (SAMs) both of a short rigid thiol, cysteamine and of a longer structurally flexible thiol, omega-amino-octanethiol (AOT). hSO on cysteamine SAMs displays a well-defined pair of voltammetric peaks around -0.207 V vs. SCE in the absence of sulfite substrate, but no electrocatalysis. hSO on AOT SAMs displays well-defined electrocatalysis, but only "fair" quality voltammetry in the absence of sulfite. We recorded next in situ scanning tunnelling spectroscopy (STS) of hSO on AOT modified Au(111)-electrodes, disclosing, a 2-5 \% surface coverage of strong molecular scale contrasts, assigned to single hSO molecules, notably with no contrast difference in the absence and presence of sulfite. In situ STS corroborated this observation with a sigmoidal tunnelling current/overpotential correlation.},
  language  = {en}
}
@article{TadjoungWaffoMitrovaTiedemannetal.2021,
  author    = {Tadjoung Waffo, Armel Franklin and Mitrova, Biljana and Tiedemann, Kim and Iobbi-Nivol, Chantal and Leimk{\"u}hler, Silke and Wollenberger, Ulla},
  title     = {Electrochemical trimethylamine n-oxide biosensor with enzyme-based oxygen-scavenging membrane for long-term operation under ambient air},
  series = {Biosensors : open access journal},
  volume    = {11},
  journal   = {Biosensors : open access journal},
  number    = {4},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2079-6374},
  doi       = {10.3390/bios11040098},
  pages     = {17},
  year      = {2021},
  abstract  = {An amperometric trimethylamine N-oxide (TMAO) biosensor is reported, where TMAO reductase (TorA) and glucose oxidase (GOD) and catalase (Cat) were immobilized on the electrode surface, enabling measurements of mediated enzymatic TMAO reduction at low potential under ambient air conditions. The oxygen anti-interference membrane composed of GOD, Cat and polyvinyl alcohol (PVA) hydrogel, together with glucose concentration, was optimized until the O-2 reduction current of a Clark-type electrode was completely suppressed for at least 3 h. For the preparation of the TMAO biosensor, Escherichia coli TorA was purified under anaerobic conditions and immobilized on the surface of a carbon electrode and covered by the optimized O-2 scavenging membrane. The TMAO sensor operates at a potential of -0.8 V vs. Ag/AgCl (1 M KCl), where the reduction of methylviologen (MV) is recorded. The sensor signal depends linearly on TMAO concentrations between 2 mu M and 15 mM, with a sensitivity of 2.75 +/- 1.7 mu A/mM. The developed biosensor is characterized by a response time of about 33 s and an operational stability over 3 weeks. Furthermore, measurements of TMAO concentration were performed in 10\% human serum, where the lowest detectable concentration is of 10 mu M TMAO.},
  language  = {en}
}
@article{LaunDuffusWahlefeldetal.2022,
  author    = {Laun, Konstantin and Duffus, Benjamin R. and Wahlefeld, Stefan and Katz, Sagie and Belger, Dennis Heinz and Hildebrandt, Peter and Mroginski, Maria Andrea and Leimk{\"u}hler, Silke and Zebger, Ingo},
  title     = {Infrared spectroscopy flucidates the inhibitor binding sites in a metal-dependent formate dehydrogenase},
  series = {Chemistry - a European journal},
  journal   = {Chemistry - a European journal},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {0947-6539},
  doi       = {10.1002/chem.202201091},
  pages     = {8},
  year      = {2022},
  abstract  = {Biological carbon dioxide (CO2) reduction is an important step by which organisms form valuable energy-richer molecules required for further metabolic processes. The Mo-dependent formate dehydrogenase (FDH) from Rhodobacter capsulatus catalyzes reversible formate oxidation to CO2 at a bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor. To elucidate potential substrate binding sites relevant for the mechanism, we studied herein the interaction with the inhibitory molecules azide and cyanate, which are isoelectronic to CO2 and charged as formate. We employed infrared (IR) spectroscopy in combination with density functional theory (DFT) and inhibition kinetics. One distinct inhibitory molecule was found to bind to either a non-competitive or a competitive binding site in the secondary coordination sphere of the active site. Site-directed mutagenesis of key amino acid residues in the vicinity of the bis-MGD cofactor revealed changes in both non-competitive and competitive binding, whereby the inhibitor is in case of the latter interaction presumably bound between the cofactor and the adjacent Arg587.},
  language  = {en}
}
@article{StrippDuffusFourmondetal.2022,
  author    = {Stripp, Sven T. and Duffus, Benjamin R. and Fourmond, Vincent and Leger, Christophe and Leimk{\"u}hler, Silke and Hirota, Shun and Hu, Yilin and Jasniewski, Andrew and Ogata, Hideaki and Ribbe, Markus W.},
  title     = {Second and outer coordination sphere effects in nitrogenase, hydrogenase, formate dehydrogenase, and CO dehydrogenase},
  series = {Chemical reviews : CR},
  volume    = {122},
  journal   = {Chemical reviews : CR},
  number    = {14},
  publisher = {American Chemical Society},
  address   = {Washington, DC},
  issn      = {0009-2665},
  doi       = {10.1021/acs.chemrev.1c00914},
  pages     = {11900 -- 11973},
  year      = {2022},
  abstract  = {Gases like H-2, N-2, CO2, and CO are increasingly recognized as critical feedstock in "green" energy conversion and as sources of nitrogen and carbon for the agricultural and chemical sectors. However, the industrial transformation of N-2, CO2, and CO and the production of H-2 require significant energy input, which renders processes like steam reforming and the Haber-Bosch reaction economically and environmentally unviable. Nature, on the other hand, performs similar tasks efficiently at ambient temperature and pressure, exploiting gas-processing metalloenzymes (GPMs) that bind low-valent metal cofactors based on iron, nickel, molybdenum, tungsten, and sulfur. Such systems are studied to understand the biocatalytic principles of gas conversion including N-2 fixation by nitrogenase and H-2 production by hydrogenase as well as CO2 and CO conversion by formate dehydrogenase, carbon monoxide dehydrogenase, and nitrogenase. In this review, we emphasize the importance of the cofactor/protein interface, discussing how second and outer coordination sphere effects determine, modulate, and optimize the catalytic activity of GPMs. These may comprise ionic interactions in the second coordination sphere that shape the electron density distribution across the cofactor, hydrogen bonding changes, and allosteric effects. In the outer coordination sphere, proton transfer and electron transfer are discussed, alongside the role of hydrophobic substrate channels and protein structural changes. Combining the information gained from structural biology, enzyme kinetics, and various spectroscopic techniques, we aim toward a comprehensive understanding of catalysis beyond the first coordination sphere.},
  language  = {en}
}
@article{FujiharaZhangJacksonetal.2022,
  author    = {Fujihara, Kenji M. and Zhang, Bonnie Z. and Jackson, Thomas D. and Ogunkola, Moses and Nijagal, Brunda and Milne, Julia V. and Sallman, David A. and Ang, Ching-Seng and Nikolic, Iva and Kearney, Conor J. and Hogg, Simon J. and Cabalag, Carlos S. and Sutton, Vivien R. and Watt, Sally and Fujihara, Asuka T. and Trapani, Joseph A. and Simpson, Kaylene J. and Stojanovski, Diana and Leimk{\"u}hler, Silke and Haupt, Sue and Phillips, Wayne A. and Clemons, Nicholas J.},
  title     = {Eprenetapopt triggers ferroptosis, inhibits NFS1 cysteine desulfurase, and synergizes with serine and glycine dietary restriction},
  series = {Science Advances},
  volume    = {8},
  journal   = {Science Advances},
  number    = {37},
  publisher = {American Assoc. for the Advancement of Science},
  address   = {Washington},
  issn      = {2375-2548},
  doi       = {10.1126/sciadv.abm9427},
  pages     = {13},
  year      = {2022},
  abstract  = {The mechanism of action of eprenetapopt (APR-246, PRIMA-1MET) as an anticancer agent remains unresolved, al-though the clinical development of eprenetapopt focuses on its reported mechanism of action as a mutant-p53 reactivator. Using unbiased approaches, this study demonstrates that eprenetapopt depletes cellular antioxidant glutathione levels by increasing its turnover, triggering a nonapoptotic, iron-dependent form of cell death known as ferroptosis. Deficiency in genes responsible for supplying cancer cells with the substrates for de novo glutathione synthesis (SLC7A11, SHMT2, and MTHFD1L), as well as the enzymes required to synthesize glutathione (GCLC and GCLM), augments the activity of eprenetapopt. Eprenetapopt also inhibits iron-sulfur cluster biogenesis by limit-ing the cysteine desulfurase activity of NFS1, which potentiates ferroptosis and may restrict cellular proliferation. The combination of eprenetapopt with dietary serine and glycine restriction synergizes to inhibit esophageal xenograft tumor growth. These findings reframe the canonical view of eprenetapopt from a mutant-p53 reactivator to a ferroptosis inducer.},
  language  = {en}
}
@article{DeSousaMotaDinizCoelhoetal.2021,
  author    = {De Sousa Mota, Cristiano and Diniz, Ana and Coelho, Catarina and Santos-Silva, Teresa and Esmaeeli Moghaddam Tabalvandani, Mariam and Leimk{\"u}hler, Silke and Cabrita, Eurico J. and Marcelo, Filipa and Rom{\~a}o, Maria Jo{\~a}o},
  title     = {Interrogating the inhibition mechanisms of human aldehyde oxidase by X-ray crystallography and NMR spectroscopy},
  series = {Journal of medicinal chemistry / American Chemical Society},
  volume    = {64},
  journal   = {Journal of medicinal chemistry / American Chemical Society},
  number    = {17},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0022-2623},
  doi       = {10.1021/acs.jmedchem.1c01125},
  pages     = {13025 -- 13037},
  year      = {2021},
  abstract  = {Human aldehyde oxidase (hAOX1) is mainly present in the liver and has an emerging role in drug metabolism, since it accepts a wide range of molecules as substrates and inhibitors. Herein, we employed an integrative approach by combining NMR, X-ray crystallography, and enzyme inhibition kinetics to understand the inhibition modes of three hAOX1 inhibitors-thioridazine, benzamidine, and raloxifene. These integrative data indicate that thioridazine is a noncompetitive inhibitor, while benzamidine presents a mixed type of inhibition. Additionally, we describe the first crystal structure of hAOX1 in complex with raloxifene. Raloxifene binds tightly at the entrance of the substrate tunnel, stabilizing the flexible entrance gates and elucidating an unusual substrate-dependent mechanism of inhibition with potential impact on drug-drug interactions. This study can be considered as a proof-of-concept for an efficient experimental screening of prospective substrates and inhibitors of hAOX1 relevant in drug discovery.},
  language  = {en}
}
@article{Leimkuehler2021,
  author    = {Leimk{\"u}hler, Silke},
  title     = {Transition metals in catalysis},
  series = {Inorganics : open access journal},
  volume    = {9},
  journal   = {Inorganics : open access journal},
  number    = {1},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2304-6740},
  doi       = {10.3390/inorganics9010006},
  pages     = {2},
  year      = {2021},
  language  = {en}
}
@article{DuffusSchrapersSchuthetal.2020,
  author    = {Duffus, Benjamin R. and Schrapers, Peer and Schuth, Nils and Mebs, Stefan and Dau, Holger and Leimk{\"u}hler, Silke and Haumann, Michael},
  title     = {Anion binding and oxidative modification at the molybdenum cofactor of formate dehydrogenase from Rhodobacter capsulatus studied by X-ray absorption spectroscopy},
  series = {Inorganic chemistry},
  volume    = {59},
  journal   = {Inorganic chemistry},
  number    = {1},
  publisher = {American Chemical Society},
  address   = {Washington, DC},
  issn      = {0020-1669},
  doi       = {10.1021/acs.inorgchem.9b01613},
  pages     = {214 -- 225},
  year      = {2020},
  abstract  = {Formate dehydrogenase (FDH) enzymes are versatile catalysts for CO2 conversion. The FDH from Rhodobacter capsulatus contains a molybdenum cofactor with the dithiolene functions of two pyranopterin guanine dinucleotide molecules, a conserved cysteine, and a sulfido group bound at Mo(VI). In this study, we focused on metal oxidation state and coordination changes in response to exposure to O-2, inhibitory anions, and redox agents using X-ray absorption spectroscopy (XAS) at the Mo K-edge. Differences in the oxidative modification of the bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor relative to samples prepared aerobically without inhibitor, such as variations in the relative numbers of sulfido (Mo=S) and oxo (Mo=O) bonds, were observed in the presence of azide (N-3(-)) or cyanate (OCN-). Azide provided best protection against O-2, resulting in a quantitatively sulfurated cofactor with a displaced cysteine ligand and optimized formate oxidation activity. Replacement of the cysteine ligand by a formate (HCO2-) ligand at the molybdenum in active enzyme is compatible with our XAS data. Cyanide (CN-) inactivated the enzyme by replacing the sulfido ligand at Mo(VI) with an oxo ligand. Evidence that the sulfido group may become protonated upon molybdenum reduction was obtained. Our results emphasize the role of coordination flexibility at the molybdenum center during inhibitory and catalytic processes of FDH enzymes.},
  language  = {en}
}
@article{MendelHercherZupoketal.2020,
  author    = {Mendel, Ralf R. and Hercher, Thomas W. and Zupok, Arkadiusz and Hasnat, Muhammad Abrar and Leimk{\"u}hler, Silke},
  title     = {The requirement of inorganic Fe-S clusters for the biosynthesis of the organometallic molybdenum cofactor},
  series = {Inorganics : open access journal},
  volume    = {8},
  journal   = {Inorganics : open access journal},
  number    = {7},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2304-6740},
  doi       = {10.3390/inorganics8070043},
  pages     = {23},
  year      = {2020},
  abstract  = {Iron-sulfur (Fe-S) clusters are essential protein cofactors. In enzymes, they are present either in the rhombic [2Fe-2S] or the cubic [4Fe-4S] form, where they are involved in catalysis and electron transfer and in the biosynthesis of metal-containing prosthetic groups like the molybdenum cofactor (Moco). Here, we give an overview of the assembly of Fe-S clusters in bacteria and humans and present their connection to the Moco biosynthesis pathway. In all organisms, Fe-S cluster assembly starts with the abstraction of sulfur froml-cysteine and its transfer to a scaffold protein. After formation, Fe-S clusters are transferred to carrier proteins that insert them into recipient apo-proteins. In eukaryotes like humans and plants, Fe-S cluster assembly takes place both in mitochondria and in the cytosol. Both Moco biosynthesis and Fe-S cluster assembly are highly conserved among all kingdoms of life. Moco is a tricyclic pterin compound with molybdenum coordinated through its unique dithiolene group. Moco biosynthesis begins in the mitochondria in a Fe-S cluster dependent step involving radical/S-adenosylmethionine (SAM) chemistry. An intermediate is transferred to the cytosol where the dithiolene group is formed, to which molybdenum is finally added. Further connections between Fe-S cluster assembly and Moco biosynthesis are discussed in detail.},
  language  = {en}
}
@misc{OgunkolaGuiraudieCaprazFeronetal.2023,
  author    = {Ogunkola, Moses Olalekan and Guiraudie-Capraz, Gaelle and F{\´e}ron, Fran{\c{c}}ois and Leimk{\"u}hler, Silke},
  title     = {The Human Mercaptopyruvate Sulfurtransferase TUM1 Is Involved in Moco Biosynthesis, Cytosolic tRNA Thiolation and Cellular Bioenergetics in Human Embryonic Kidney Cells},
  series = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  number    = {1307},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-57958},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-579580},
  pages     = {23},
  year      = {2023},
  abstract  = {Sulfur is an important element that is incorporated into many biomolecules in humans. The incorporation and transfer of sulfur into biomolecules is, however, facilitated by a series of different sulfurtransferases. Among these sulfurtransferases is the human mercaptopyruvate sulfurtransferase (MPST) also designated as tRNA thiouridine modification protein (TUM1). The role of the human TUM1 protein has been suggested in a wide range of physiological processes in the cell among which are but not limited to involvement in Molybdenum cofactor (Moco) biosynthesis, cytosolic tRNA thiolation and generation of H2S as signaling molecule both in mitochondria and the cytosol. Previous interaction studies showed that TUM1 interacts with the L-cysteine desulfurase NFS1 and the Molybdenum cofactor biosynthesis protein 3 (MOCS3). Here, we show the roles of TUM1 in human cells using CRISPR/Cas9 genetically modified Human Embryonic Kidney cells. Here, we show that TUM1 is involved in the sulfur transfer for Molybdenum cofactor synthesis and tRNA thiomodification by spectrophotometric measurement of the activity of sulfite oxidase and liquid chromatography quantification of the level of sulfur-modified tRNA. Further, we show that TUM1 has a role in hydrogen sulfide production and cellular bioenergetics.},
  language  = {en}
}
@article{OgunkolaGuiraudieCaprazFeronetal.2023,
  author    = {Ogunkola, Moses Olalekan and Guiraudie-Capraz, Gaelle and F{\´e}ron, Fran{\c{c}}ois and Leimk{\"u}hler, Silke},
  title     = {The Human Mercaptopyruvate Sulfurtransferase TUM1 Is Involved in Moco Biosynthesis, Cytosolic tRNA Thiolation and Cellular Bioenergetics in Human Embryonic Kidney Cells},
  series = {Biomolecules},
  volume    = {13},
  journal   = {Biomolecules},
  edition   = {1},
  publisher = {MDPI},
  address   = {Basel, Schweiz},
  issn      = {2218-273X},
  doi       = {10.3390/biom13010144},
  pages     = {1 -- 23},
  year      = {2023},
  abstract  = {Sulfur is an important element that is incorporated into many biomolecules in humans. The incorporation and transfer of sulfur into biomolecules is, however, facilitated by a series of different sulfurtransferases. Among these sulfurtransferases is the human mercaptopyruvate sulfurtransferase (MPST) also designated as tRNA thiouridine modification protein (TUM1). The role of the human TUM1 protein has been suggested in a wide range of physiological processes in the cell among which are but not limited to involvement in Molybdenum cofactor (Moco) biosynthesis, cytosolic tRNA thiolation and generation of H2S as signaling molecule both in mitochondria and the cytosol. Previous interaction studies showed that TUM1 interacts with the L-cysteine desulfurase NFS1 and the Molybdenum cofactor biosynthesis protein 3 (MOCS3). Here, we show the roles of TUM1 in human cells using CRISPR/Cas9 genetically modified Human Embryonic Kidney cells. Here, we show that TUM1 is involved in the sulfur transfer for Molybdenum cofactor synthesis and tRNA thiomodification by spectrophotometric measurement of the activity of sulfite oxidase and liquid chromatography quantification of the level of sulfur-modified tRNA. Further, we show that TUM1 has a role in hydrogen sulfide production and cellular bioenergetics.},
  language  = {en}
}
@article{GarridoLeimkuehler2021,
  author    = {Garrido, Claudia and Leimk{\"u}hler, Silke},
  title     = {The inactivation of human aldehyde oxidase 1 by hydrogen peroxide and superoxide},
  series = {Drug metabolism and disposition / American Society for Pharmacology and Experimental Therapeutics},
  volume    = {49},
  journal   = {Drug metabolism and disposition / American Society for Pharmacology and Experimental Therapeutics},
  number    = {9},
  publisher = {American Society for Pharmacology and Experimental Therapeutics},
  address   = {Bethesda},
  issn      = {1521-009X},
  doi       = {10.1124/dmd.121.000549},
  pages     = {729 -- 735},
  year      = {2021},
  abstract  = {Mammalian aldehyde oxidases (AOX) are molybdo-flavoenzymes of pharmacological and pathophysiologic relevance that are involved in phase I drug metabolism and, as a product of their enzymatic activity, are also involved in the generation of reactive oxygen species. So far, the physiologic role of aldehyde oxidase 1 in the human body remains unknown. The human enzyme hAOX1 is characterized by a broad substrate specificity, oxidizing aromatic/aliphatic aldehydes into their corresponding carboxylic acids, and hydroxylating various heteroaromatic rings. The enzyme uses oxygen as terminal electron acceptor to produce hydrogen peroxide and superoxide during turnover. Since hAOX1 and, in particular, some natural variants produce not only H2O2 but also high amounts of superoxide, we investigated the effect of both ROS molecules on the enzymatic activity of hAOX1 in more detail. We compared hAOX1 to the high-O-2(.-)-producing natural variant L438V for their time-dependent inactivation with H2O2/O-2(.-) during substrate turnover. We show that the inactivation of the hAOX1 wild-type enzyme is mainly based on the production of hydrogen peroxide, whereas for the variant L438V, both hydrogen peroxide and superoxide contribute to the time-dependent inactivation of the enzyme during turnover. Further, the level of inactivation was revealed to be substrate-dependent: using substrates with higher turnover numbers resulted in a faster inactivation of the enzymes. Analysis of the inactivation site of the enzyme identified a loss of the terminal sulfido ligand at the molybdenum active site by the produced ROS during turnover.},
  language  = {en}
}
@article{YildizLeimkuehler2021,
  author    = {Yildiz, Tugba and Leimk{\"u}hler, Silke},
  title     = {TusA is a versatile protein that links translation efficiency to cell division in Escherichia coli},
  series = {Journal of bacteriology},
  volume    = {203},
  journal   = {Journal of bacteriology},
  number    = {7},
  publisher = {American Society for Microbiology},
  address   = {Washington},
  issn      = {1098-5530},
  doi       = {10.1128/JB.00659-20},
  pages     = {20},
  year      = {2021},
  abstract  = {To enable accurate and efficient translation, sulfur modifications are introduced posttranscriptionally into nucleosides in tRNAs. The biosynthesis of tRNA sulfur modifications involves unique sulfur trafficking systems for the incorporation of sulfur atoms in different nucleosides of tRNA. One of the proteins that is involved in inserting the sulfur for 5-methylaminomethyl-2-thiouridine (mnm(5)s(2)U34) modifications in tRNAs is the TusA protein. TusA, however, is a versatile protein that is also involved in numerous other cellular pathways. Despite its role as a sulfur transfer protein for the 2-thiouridine formation in tRNA, a fundamental role of TusA in the general physiology of Escherichia coli has also been discovered. Poor viability, a defect in cell division, and a filamentous cell morphology have been described previously for tusA-deficient cells. In this report, we aimed to dissect the role of TusA for cell viability. We were able to show that the lack of the thiolation status of wobble uridine (U-34) nucleotides present on Lys, Gln, or Glu in tRNAs has a major consequence on the translation efficiency of proteins; among the affected targets are the proteins RpoS and Fis. Both proteins are major regulatory factors, and the deregulation of their abundance consequently has a major effect on the cellular regulatory network, with one consequence being a defect in cell division by regulating the FtsZ ring formation. <br /> IMPORTANCE More than 100 different modifications are found in RNAs. One of these modifications is the mnm(5)s(2)U modification at the wobble position 34 of tRNAs for Lys, Gln, and Glu. The functional significance of U34 modifications is substantial since it restricts the conformational flexibility of the anticodon, thus providing translational fidelity. We show that in an Escherichia coli TusA mutant strain, involved in sulfur transfer for the mnm(5)s(2)U34 thio modifications, the translation efficiency of RpoS and Fis, two major cellular regulatory proteins, is altered. Therefore, in addition to the transcriptional regulation and the factors that influence protein stability, tRNA modifications that ensure the translational efficiency provide an additional crucial regulatory factor for protein synthesis.},
  language  = {en}
}
@article{HasnatZupokOlasApeltetal.2021,
  author    = {Hasnat, Muhammad Abrar and Zupok, Arkadiusz and Olas-Apelt, Justyna Jadwiga and M{\"u}ller-R{\"o}ber, Bernd and Leimk{\"u}hler, Silke},
  title     = {A-type carrier proteins are involved in [4Fe-4S] cluster insertion into the radical S-adenosylmethionine protein MoaA for the synthesis of active molybdoenzymes},
  series = {Journal of bacteriology},
  volume    = {203},
  journal   = {Journal of bacteriology},
  number    = {12},
  publisher = {American Society for Microbiology},
  address   = {Washington},
  issn      = {1098-5530},
  doi       = {10.1128/JB.00086-21},
  pages     = {20},
  year      = {2021},
  abstract  = {Iron sulfur (Fe-S) clusters are important biological cofactors present in proteins with crucial biological functions, from photosynthesis to DNA repair, gene expression, and bioenergetic processes. For the insertion of Fe-S clusters into proteins, A-type carrier proteins have been identified. So far, three of them have been characterized in detail in Escherichia coli, namely, IscA, SufA, and ErpA, which were shown to partially replace each other in their roles in [4Fe-4S] cluster insertion into specific target proteins. To further expand the knowledge of [4Fe-4S] cluster insertion into proteins, we analyzed the complex Fe-S cluster-dependent network for the synthesis of the molybdenum cofactor (Moco) and the expression of genes encoding nitrate reductase in E. coli. Our studies include the identification of the A-type carrier proteins ErpA and IscA, involved in [4Fe-4S] cluster insertion into the radical Sadenosyl-methionine (SAM) enzyme MoaA. We show that ErpA and IscA can partially replace each other in their role to provide [4Fe-4S] clusters for MoaA. Since most genes expressing molybdoenzymes are regulated by the transcriptional regulator for fumarate and nitrate reduction (FNR) under anaerobic conditions, we also identified the proteins that are crucial to obtain an active FNR under conditions of nitrate respiration. We show that ErpA is essential for the FNR-dependent expression of the narGHJI operon, a role that cannot be compensated by IscA under the growth conditions tested. SufA does not appear to have a role in Fe-S cluster insertion into MoaA or FNR under anaerobic growth employing nitrate respiration, based on the low level of gene expression. <br /> IMPORTANCE Understanding the assembly of iron-sulfur (Fe-S) proteins is relevant to many fields, including nitrogen fixation, photosynthesis, bioenergetics, and gene regulation. Remaining critical gaps in our knowledge include how Fe-S clusters are transferred to their target proteins and how the specificity in this process is achieved, since different forms of Fe-S clusters need to be delivered to structurally highly diverse target proteins. Numerous Fe-S carrier proteins have been identified in prokaryotes like Escherichia coli, including ErpA, IscA, SufA, and NfuA. In addition, the diverse Fe-S cluster delivery proteins and their target proteins underlie a complex regulatory network of expression, to ensure that both proteins are synthesized under particular growth conditions.},
  language  = {en}
}
@misc{Leimkuehler2020,
  author    = {Leimk{\"u}hler, Silke},
  title     = {The biosynthesis of the molybdenum cofactors in Escherichia coli},
  series = {Environmental microbiology},
  volume    = {22},
  journal   = {Environmental microbiology},
  number    = {6},
  publisher = {Wiley},
  address   = {Hoboken},
  issn      = {1462-2912},
  doi       = {10.1111/1462-2920.15003},
  pages     = {2007 -- 2026},
  year      = {2020},
  abstract  = {The biosynthesis of the molybdenum cofactor (Moco) is highly conserved among all kingdoms of life. In all molybdoenzymes containing Moco, the molybdenum atom is coordinated to a dithiolene group present in the pterin-based 6-alkyl side chain of molybdopterin (MPT). In general, the biosynthesis of Moco can be divided into four steps in in bacteria: (i) the starting point is the formation of the cyclic pyranopterin monophosphate (cPMP) from 5 '-GTP, (ii) in the second step the two sulfur atoms are inserted into cPMP leading to the formation of MPT, (iii) in the third step the molybdenum atom is inserted into MPT to form Moco and (iv) in the fourth step bis-Mo-MPT is formed and an additional modification of Moco is possible with the attachment of a nucleotide (CMP or GMP) to the phosphate group of MPT, forming the dinucleotide variants of Moco. This review presents an update on the well-characterized Moco biosynthesis in the model organism Escherichia coli including novel discoveries from the recent years.},
  language  = {en}
}
@article{NishinoOkamotoLeimkuehler2017,
  author    = {Nishino, Takeshi and Okamoto, Ken and Leimk{\"u}hler, Silke},
  title     = {Enzymes of the Xanthine Oxidase Family},
  series = {Molybdenum and tungsten enzymes : biochemistry},
  volume    = {5},
  journal   = {Molybdenum and tungsten enzymes : biochemistry},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  isbn      = {978-1-78262-391-5},
  doi       = {10.1039/9781782623915-00192},
  pages     = {192 -- 239},
  year      = {2017},
  abstract  = {Enzymes from the xanthine oxidase (XO) family of molybdenum enzymes are generally, with some exceptions, molybdenum iron-sulfur flavin hydroxylases. Mammalian xanthine oxidoreductase and aldehyde oxidase were among the first enzymes to be studied in detail more than 100 years ago and, surprisingly, they continue to be thoroughly studied in molecular detail with many open and unresolved questions remaining. Enzymes of the XO family are characterized by a molybdenum cofactor (Moco) active site with a MoVIOS(OH) ligand sphere where substrate hydroxylation of either aromatic or aliphatic carbon centers is catalyzed. During the reaction, electrons are transferred to the oxidizing substrate, most commonly O2 or NAD+, which react at the FAD site.},
  language  = {en}
}
@article{LeimkuehlerLemaireIobbiNivol2017,
  author    = {Leimk{\"u}hler, Silke and Lemaire, Olivier N. and Iobbi-Nivol, Chantal},
  title     = {Bacterial Molybdoenzymes},
  series = {Molybdenum and tungsten enzymes : biochemistry},
  volume    = {5},
  journal   = {Molybdenum and tungsten enzymes : biochemistry},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  isbn      = {978-1-78262-391-5},
  doi       = {10.1039/9781782623915-00117},
  pages     = {117 -- 142},
  year      = {2017},
  abstract  = {The biogenesis of molybdoenzymes is a cytoplasmic event requiring both the folded apoenzymes and the matured molybdenum cofactor. The structure and the complexity of the molybdenum cofactor varies in each molybdoenzyme family and consequently different accessory proteins are required for the maturation of the respective enzymes. Thus, for enzymes of both the DMSO reductase and xanthine oxidase families, specific chaperones exist which are dedicated to increase the stability and the folding of specific members of each family. In this review, we describe the role of these chaperones for molybdoenzyme maturation. We present a model which describes step by step the mechanism of the maturation of representative molybdoenzymes from each family.},
  language  = {en}
}
@article{LeimkuehlerMendel2017,
  author    = {Leimk{\"u}hler, Silke and Mendel, Ralf-Rainer},
  title     = {Molybdenum Cofactor Biosynthesis},
  series = {Molybdenum and tungsten enzymes: biochemistry},
  volume    = {5},
  journal   = {Molybdenum and tungsten enzymes: biochemistry},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  isbn      = {978-1-78262-391-5},
  doi       = {10.1039/9781782623915},
  pages     = {100 -- 116},
  year      = {2017},
  abstract  = {The biosynthesis of the molybdenum cofactor (Moco) is highly conserved among all kingdoms of life. In all molybdoenzymes with the exception of nitrogenase, the molybdenum atom is coordinated to a dithiolene group present in the pterin-based 6-alkyl side chain of molybdopterin (MPT). In general, the biosynthesis of Moco can be divided into three steps in eukaryotes, and four steps in bacteria and archaea: (i) the starting point is the formation of the cyclic pyranopterin monophosphate (cPMP) from 5′GTP, (ii) in the second step the two sulfur molecules are inserted into cPMP leading to the formation of MPT, (iii) in the third step the molybdenum atom is inserted into molybdopterin to form Moco and (iv) additional modification of Moco occurs in bacteria and archaea with the attachment of a nucleotide (CMP or GMP) to the phosphate group of MPT, forming the dinucleotide variants of Moco. This review will focus on the biosynthesis of Moco in bacteria, humans and plants.},
  language  = {en}
}
@misc{TiedemannIobbiNivolLeimkuehler2022,
  author    = {Tiedemann, Kim and Iobbi-Nivol, Chantal and Leimk{\"u}hler, Silke},
  title     = {The Role of the Nucleotides in the Insertion of the bis-Molybdopterin Guanine Dinucleotide Cofactor into apo-Molybdoenzymes},
  series = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  publisher = {Universit{\"a}tsverlag Potsdam},
  address   = {Potsdam},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-56172},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-561728},
  pages     = {1 -- 15},
  year      = {2022},
  abstract  = {The role of the GMP nucleotides of the bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor of the DMSO reductase family has long been a subject of discussion. The recent characterization of the bis-molybdopterin (bis-Mo-MPT) cofactor present in the E. coli YdhV protein, which differs from bis-MGD solely by the absence of the nucleotides, now enables studying the role of the nucleotides of bis-MGD and bis-MPT cofactors in Moco insertion and the activity of molybdoenzymes in direct comparison. Using the well-known E. coli TMAO reductase TorA as a model enzyme for cofactor insertion, we were able to show that the GMP nucleotides of bis-MGD are crucial for the insertion of the bis-MGD cofactor into apo-TorA.},
  language  = {en}
}
@article{TiedemannIobbiNivolLeimkuehler2022,
  author    = {Tiedemann, Kim and Iobbi-Nivol, Chantal and Leimk{\"u}hler, Silke},
  title     = {The Role of the Nucleotides in the Insertion of the bis-Molybdopterin Guanine Dinucleotide Cofactor into apo-Molybdoenzymes},
  series = {Molecules},
  volume    = {27},
  journal   = {Molecules},
  edition   = {9},
  publisher = {MDPI},
  address   = {Basel, Schweiz},
  issn      = {1420-3049},
  doi       = {10.3390/molecules27092993},
  pages     = {1 -- 15},
  year      = {2022},
  abstract  = {The role of the GMP nucleotides of the bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor of the DMSO reductase family has long been a subject of discussion. The recent characterization of the bis-molybdopterin (bis-Mo-MPT) cofactor present in the E. coli YdhV protein, which differs from bis-MGD solely by the absence of the nucleotides, now enables studying the role of the nucleotides of bis-MGD and bis-MPT cofactors in Moco insertion and the activity of molybdoenzymes in direct comparison. Using the well-known E. coli TMAO reductase TorA as a model enzyme for cofactor insertion, we were able to show that the GMP nucleotides of bis-MGD are crucial for the insertion of the bis-MGD cofactor into apo-TorA.},
  language  = {en}
}
@misc{LeimkuehlerBuehningBeilschmidt2017,
  author    = {Leimk{\"u}hler, Silke and B{\"u}hning, Martin and Beilschmidt, Lena},
  title     = {Shared sulfur mobilization routes for tRNA thiolation and molybdenum cofactor biosynthesis in prokaryotes and eukaryotes},
  series = {Biomolecules},
  volume    = {7},
  journal   = {Biomolecules},
  number    = {1},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2218-273X},
  doi       = {10.3390/biom7010005},
  pages     = {20},
  year      = {2017},
  abstract  = {Modifications of transfer RNA (tRNA) have been shown to play critical roles in the biogenesis, metabolism, structural stability and function of RNA molecules, and the specific modifications of nucleobases with sulfur atoms in tRNA are present in pro- and eukaryotes. Here, especially the thiomodifications xm(5)s(2)U at the wobble position 34 in tRNAs for Lys, Gln and Glu, were suggested to have an important role during the translation process by ensuring accurate deciphering of the genetic code and by stabilization of the tRNA structure. The trafficking and delivery of sulfur nucleosides is a complex process carried out by sulfur relay systems involving numerous proteins, which not only deliver sulfur to the specific tRNAs but also to other sulfur-containing molecules including iron-sulfur clusters, thiamin, biotin, lipoic acid and molybdopterin (MPT). Among the biosynthesis of these sulfur-containing molecules, the biosynthesis of the molybdenum cofactor (Moco) and the synthesis of thio-modified tRNAs in particular show a surprising link by sharing protein components for sulfur mobilization in pro- and eukaryotes.},
  language  = {en}
}
@article{FriemelMareljaLietal.2017,
  author    = {Friemel, Martin and Marelja, Zvonimir and Li, Kuanyu and Leimk{\"u}hler, Silke},
  title     = {The N-Terminus of Iron-Sulfur Cluster Assembly Factor ISD11 Is Crucial for Subcellular Targeting and Interaction with L-Cysteine Desulfurase NFS1},
  series = {Biochemistry},
  volume    = {56},
  journal   = {Biochemistry},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.6b01239},
  pages     = {1797 -- 1808},
  year      = {2017},
  abstract  = {Assembly of iron sulfur (FeS) clusters is an important process in living cells. The initial sulfur mobilization step for FeS cluster biosynthesis is catalyzed by L-cysteine desulfurase NFS1, a reaction that is localized in mitochondria in humans. In humans, the function of NFS1 depends on the ISD11 protein, which is required to stabilize its structure. The NFS1/ISD11 complex further interacts with scaffold protein ISCU and regulator protein frataxin, thereby forming a quaternary complex for FeS cluster formation. It has been suggested that the role of ISD11 is not restricted to its role in stabilizing the structure of NFS1, because studies of single-amino acid variants of ISD11 additionally demonstrated its importance for the correct assembly of the quaternary complex. In this study, we are focusing on the N-terminal region of ISD11 to determine the role of N-terminal amino acids in the formation of the complex with NFS1 and to reveal the mitochondria) targeting sequence for subcellular localization. Our in vitro studies with the purified proteins and in vivo studies in a cellular system show that the first 10 N-terminal amino acids of ISD11 are indispensable for the activity of NFS1 and especially the conserved "LYR" motif is essential for the role of ISD11 in forming a stable and active complex with NFS1.},
  language  = {en}
}
@misc{RomaoCoelhoSantosSilvaetal.2017,
  author    = {Romao, Maria Joao and Coelho, Catarina and Santos-Silva, Teresa and Foti, Alessandro and Terao, Mineko and Garattini, Enrico and Leimk{\"u}hler, Silke},
  title     = {Structural basis for the role of mammalian aldehyde oxidases in the metabolism of drugs and xenobiotics},
  series = {Current Opinion in Chemical Biology},
  volume    = {37},
  journal   = {Current Opinion in Chemical Biology},
  publisher = {Elsevier},
  address   = {Oxford},
  issn      = {1367-5931},
  doi       = {10.1016/j.cbpa.2017.01.005},
  pages     = {39 -- 47},
  year      = {2017},
  abstract  = {Aldehyde oxidases (AOXs) are molybdo-flavoenzymes characterized by broad substrate specificity, oxidizing aromatic/aliphatic aldehydes into the corresponding carboxylic acids and hydroxylating various heteroaromatic rings. Mammals are characterized by a complement of species specific AOX isoenzymes, that varies from one in humans (AOX1) to four in rodents (AOX1, AOX2, AOX3 and AOX4). The physiological function of mammalian AOX isoenzymes is unknown, although human AOX1 is an emerging enzyme in phase-I drug metabolism. Indeed, the number of therapeutic molecules under development which act as AOX substrates is increasing. The recent crystallization and structure determination of human AOX1 as well as mouse AOX3 has brought new insights into the mechanisms underlying substrate/inhibitor binding as well as the catalytic activity of this class of enzymes.},
  language  = {en}
}
@article{BuehningVallerianiLeimkuehler2017,
  author    = {B{\"u}hning, Martin and Valleriani, Angelo and Leimk{\"u}hler, Silke},
  title     = {The role of SufS is restricted to Fe-S cluster biosynthesis in escherichia coli},
  series = {Biochemistry},
  volume    = {56},
  journal   = {Biochemistry},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.7b00040},
  pages     = {1987 -- 2000},
  year      = {2017},
  abstract  = {In Escherichia coli, two different systems that are important for the coordinate formation of Fe-S clusters have been identified, namely, the ISC and SUF systems. The ISC system is the housekeeping Fe-S machinery, which provides Fe-S clusters for numerous cellular proteins. The IscS protein of this system was additionally revealed to be the primary sulfur donor for several sulfur-containing molecules with important biological functions, among which are the molybdenum cofactor (Moco) and thiolated nucleosides in tRNA. Here, we show that deletion of central components of the ISC system in addition to IscS leads to an overall decrease in Fe-S cluster enzyme and molybdoenzyme activity in addition to a decrease in the number of Fe-S-dependent thiomodifications of tRNA, based on the fact that some proteins involved in Moco biosynthesis and tRNA thiolation are Fe-S-dependent. Complementation of the ISC deficient strains with the suf operon restored the activity of Fe-S-containing proteins, including the MoaA protein, which is involved in the conversion of 5′GTP to cyclic pyranopterin monophosphate in the fist step of Moco biosynthesis. While both systems share a high degree of similarity, we show that the function of their respective l-cysteine desulfurase IscS or SufS is specific for each cellular pathway. It is revealed that SufS cannot play the role of IscS in sulfur transfer for the formation of 2-thiouridine, 4-thiouridine, or the dithiolene group of molybdopterin, being unable to interact with TusA or ThiI. The results demonstrate that the role of the SUF system is exclusively restricted to Fe-S cluster assembly in the cell.},
  language  = {en}
}
@article{ParagasHumphreysMinetal.2017,
  author    = {Paragas, Erickson M. and Humphreys, Sara C. and Min, Joshua and Joswig-Jones, Carolyn A. and Leimk{\"u}hler, Silke and Jones, Jeffrey P.},
  title     = {ecoAO},
  series = {ACS OMEGA},
  volume    = {2},
  journal   = {ACS OMEGA},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {2470-1343},
  doi       = {10.1021/acsomega.7b01054},
  pages     = {4820 -- 4827},
  year      = {2017},
  abstract  = {Although aldehyde oxidase (AO) is an important hepatic drug-metabolizing enzyme, it remains understudied and is consequently often overlooked in preclinical studies, an oversight that has resulted in the failure of multiple clinical trials. AO's preclusion to investigation stems from the following: (1) difficulties synthesizing metabolic standards due to the chemospecificity and regiospecificity of the enzyme and (2) significant inherent variability across existing in vitro systems including liver cytosol, S9 fractions, and primary hepatocytes, which lack specificity and generate discordant expression and activity profiles. Here, we describe a practical bacterial biotransformation system, ecoAO, addressing both issues simultaneously. ecoAO is a cell paste of MoCo-producing Escherichia coli strain TP1017 expressing human AO. It exhibits specific activity toward known substrates, zoniporide, 4-trans-(N,N-dimethylamino)cinnamaldehyde, O6-benzylguanine, and zaleplon; it also has utility as a biocatalyst, yielding milligram quantities of synthetically challenging metabolite standards such as 2-oxo-zoniporide. Moreover, ecoAO enables routine determination of kcat and V/K, which are essential parameters for accurate in vivo clearance predictions. Furthermore, ecoAO has potential as a preclinical in vitro screening tool for AO activity, as demonstrated by its metabolism of 3-aminoquinoline, a previously uncharacterized substrate. ecoAO promises to provide easy access to metabolites with the potential to improve pharmacokinetic clearance predictions and guide drug development.},
  language  = {en}
}
@article{KuecuekgoezeLeimkuehler2018,
  author    = {K{\"u}{\c{c}}{\"u}kg{\"o}ze, G{\"o}khan and Leimk{\"u}hler, Silke},
  title     = {Direct comparison of the four aldehyde oxidase enzymes present in mouse gives insight into their substrate specificities},
  series = {PLOS ONE},
  volume    = {13},
  journal   = {PLOS ONE},
  number    = {1},
  publisher = {Public Library of Science},
  address   = {San Fransisco},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0191819},
  pages     = {20},
  year      = {2018},
  abstract  = {Mammalian aldehyde oxidases (AOXs) are molybdo-flavoenzymes which are present in many tissues in various mammalian species, including humans and rodents. Different species contain a different number of AOX isoforms. In particular, the reasons why mammals other than humans express a multiplicity of tissue-specific AOX enzymes is unknown. In mouse, the isoforms mAOX1, mAOX3, mAOX4 and mAOX2 are present. We previously established a codon-optimized heterologous expression systems for the mAOX1-4 isoforms in Escherichia coli that gives yield to sufficient amounts of active protein for kinetic characterizations and sets the basis in this study for site-directed mutagenesis and structure-function studies. A direct and simultaneous comparison of the enzymatic properties and characteristics of the four enzymes on a larger number of substrates has never been performed. Here, thirty different structurally related aromatic, aliphatic and N-heterocyclic compounds were used as substrates, and the kinetic parameters of all four mAOX enzymes were directly compared. The results show that especially mAOX4 displays a higher substrate selectivity, while no major differences between mAOX1, mAOX2 and mAOX3 were identified. Generally, mAOX1 was the enzyme with the highest catalytic turnover for most substrates. To understand the factors that contribute to the substrate specificity of mAOX4, site-directed mutagenesis was applied to substitute amino acids in the substrate-binding funnel by the ones present in mAOX1, mAOX3, and mAOX2. An increase in activity was obtained by the amino acid exchange M1088V in the active site identified to be specific for mAOX4, to the amino acid identified in mAOX3.},
  language  = {en}
}
@article{KaufmannDuffusMitrovaetal.2018,
  author    = {Kaufmann, Hans Paul and Duffus, Benjamin R. and Mitrova, Biljana and Iobbi-Nivol, Chantal and Teutloff, Christian and Nimtz, Manfred and Jaensch, Lothar and Wollenberger, Ulla and Leimk{\"u}hler, Silke},
  title     = {Modulating the Molybdenum Coordination Sphere of Escherichia coli Trimethylamie N-Oxide Reductase},
  series = {Biochemistry},
  volume    = {57},
  journal   = {Biochemistry},
  number    = {7},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.7b01108},
  pages     = {1130 -- 1143},
  year      = {2018},
  abstract  = {The well-studied enterobacterium Escherichia coli present in the human gut can reduce trimethylamine N-oxide (TMAO) to trimethylamine during anaerobic respiration. The TMAO reductase TorA is a monomeric, bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor-containing enzyme that belongs to the dimethyl sulfoxide reductase family of molybdoenzymes. We report on a system for the in vitro reconstitution of TorA with molybdenum cofactors (Moco) from different sources. Higher TMAO reductase activities for TorA were obtained when using Moco sources containing a sulfido ligand at the molybdenum atom. For the first time, we were able to isolate functional bis-MGD from Rhodobacter capsulatus formate dehydrogenase (FDH), which remained intact in its isolated state and after insertion into apo-TorA yielded a highly active enzyme. Combined characterizations of the reconstituted TorA enzymes by electron paramagnetic resonance spectroscopy and direct electrochemistry emphasize that TorA activity can be modified by changes in the Mo coordination sphere. The combination of these results together with studies of amino acid exchanges at the active site led us to propose a novel model for binding of the substrate to the molybdenum atom of TorA.},
  language  = {en}
}
@article{KaufmannDuffusTeutloffetal.2018,
  author    = {Kaufmann, Paul and Duffus, Benjamin R. and Teutloff, Christian and Leimk{\"u}hler, Silke},
  title     = {Functional Studies on Oligotropha carboxidovorans Molybdenum-Copper CO Dehydrogenase Produced in Escherichia coli},
  series = {Biochemistry},
  volume    = {57},
  journal   = {Biochemistry},
  number    = {19},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.8b00128},
  pages     = {2889 -- 2901},
  year      = {2018},
  abstract  = {The Mo/Cu-dependent CO dehydrogenase (CODH) from Oligotropha carboxidovorans is an enzyme that is able to catalyze both the oxidation of CO to CO2 and the oxidation of H-2 to protons and electrons. Despite the close to atomic resolution structure (1.1 angstrom), significant uncertainties have remained with regard to the reaction mechanism of substrate oxidation at the unique Mo/Cu center, as well as the nature of intermediates formed during the catalytic cycle. So far, the investigation of the role of amino acids at the active site was hampered by the lack of a suitable expression system that allowed for detailed site-directed mutagenesis studies at the active site. Here, we report on the establishment of a functional heterologous expression system of O. carboxidovorans CODH in Escherichia coli. We characterize the purified enzyme in detail by a combination of kinetic and spectroscopic studies and show that it was purified in a form with characteristics comparable to those of the native enzyme purified from O. carboxidovorans. With this expression system in hand, we were for the first time able to generate active-site variants of this enzyme. Our work presents the basis for more detailed studies of the reaction mechanism for CO and H-2 oxidation of Mo/Cu-dependent CODHs in the future.},
  language  = {en}
}
@article{OttoMareljaSchoofsetal.2018,
  author    = {Otto, Nils and Marelja, Zvonimir and Schoofs, Andreas and Kranenburg, Holger and Bittern, Jonas and Yildirim, Kerem and Berh, Dimitri and Bethke, Maria and Thomas, Silke and Rode, Sandra and Risse, Benjamin and Jiang, Xiaoyi and Pankratz, Michael and Leimk{\"u}hler, Silke and Kl{\"a}mbt, Christian},
  title     = {The sulfite oxidase Shopper controls neuronal activity by regulating glutamate homeostasis in Drosophila ensheathing glia},
  series = {Nature Communications},
  volume    = {9},
  journal   = {Nature Communications},
  publisher = {Nature Publ. Group},
  address   = {London},
  issn      = {2041-1723},
  doi       = {10.1038/s41467-018-05645-z},
  pages     = {12},
  year      = {2018},
  abstract  = {Specialized glial subtypes provide support to developing and functioning neural networks. Astrocytes modulate information processing by neurotransmitter recycling and release of neuromodulatory substances, whereas ensheathing glial cells have not been associated with neuromodulatory functions yet. To decipher a possible role of ensheathing glia in neuronal information processing, we screened for glial genes required in the Drosophila central nervous system for normal locomotor behavior. Shopper encodes a mitochondrial sulfite oxidase that is specifically required in ensheathing glia to regulate head bending and peristalsis. shopper mutants show elevated sulfite levels affecting the glutamate homeostasis which then act on neuronal network function. Interestingly, human patients lacking the Shopper homolog SUOX develop neurological symptoms, including seizures. Given an enhanced expression of SUOX by oligodendrocytes, our findings might indicate that in both invertebrates and vertebrates more than one glial cell type may be involved in modulating neuronal activity.},
  language  = {en}
}
@article{SchwanholdIobbiNivolLehmannetal.2018,
  author    = {Schwanhold, Nadine and Iobbi-Nivol, Chantal and Lehmann, Angelika and Leimk{\"u}hler, Silke},
  title     = {Same but different},
  series = {PLoS one},
  volume    = {13},
  journal   = {PLoS one},
  number    = {11},
  publisher = {PLoS},
  address   = {San Fransisco},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0201935},
  pages     = {24},
  year      = {2018},
  abstract  = {The maturation of bacterial molybdoenzymes is a complex process leading to the insertion of the bulky bis-molybdopterin guanine dinucleotide (bis-MGD) cofactor into the apoenzyme. Most molybdoenzymes were shown to contain a specific chaperone for the insertion of the bis-MGD cofactor. Formate dehydrogenases (FDH) together with their molecular chaperone partner seem to display an exception to this specificity rule, since the chaperone FdhD has been proven to be involved in the maturation of all three FDH enzymes present in Escherichia colt. Multiple roles have been suggested for FdhD-like chaperones in the past, including the involvement in a sulfur transfer reaction from the L-cysteine desulfurase IscS to bis-MGD by the action of two cysteine residues present in a conserved CXXC motif of the chaperones. However, in this study we show by phylogenetic analyses that the CXXC motif is not conserved among FdhD-like chaperones. We compared in detail the FdhD-like homologues from Rhodobacter capsulatus and E. colt and show that their roles in the maturation of FDH enzymes from different subgroups can be exchanged. We reveal that bis-MGDbinding is a common characteristic of FdhD-like proteins and that the cofactor is bound with a sulfido-ligand at the molybdenum atom to the chaperone. Generally, we reveal that the cysteine residues in the motif CXXC of the chaperone are not essential for the production of active FDH enzymes.},
  language  = {en}
}
@article{BurschelDecovicNuberetal.2018,
  author    = {Burschel, Sabrina and Decovic, Doris Kreuzer and Nuber, Franziska and Stiller, Marie and Hofmann, Maud and Zupok, Arkadiusz and Siemiatkowska, Beata and Gorka, Michal Jakub and Leimk{\"u}hler, Silke and Friedrich, Thorsten},
  title     = {Iron-sulfur cluster carrier proteins involved in the assembly of Escherichia coli NADH},
  series = {Molecular microbiology},
  volume    = {111},
  journal   = {Molecular microbiology},
  number    = {1},
  publisher = {Wiley},
  address   = {Hoboken},
  issn      = {0950-382X},
  doi       = {10.1111/mmi.14137},
  pages     = {31 -- 45},
  year      = {2018},
  abstract  = {The NADH:ubiquinone oxidoreductase (respiratory complex I) is the main entry point for electrons into the Escherichia coli aerobic respiratory chain. With its sophisticated setup of 13 different subunits and 10 cofactors, it is anticipated that various chaperones are needed for its proper maturation. However, very little is known about the assembly of E. coli complex I, especially concerning the incorporation of the iron-sulfur clusters. To identify iron-sulfur cluster carrier proteins possibly involved in the process, we generated knockout strains of NfuA, BolA, YajL, Mrp, GrxD and IbaG that have been reported either to be involved in the maturation of mitochondrial complex I or to exert influence on the clusters of bacterial complex. We determined the NADH and succinate oxidase activities of membranes from the mutant strains to monitor the specificity of the individual mutations for complex I. The deletion of NfuA, BolA and Mrp led to a decreased stability and partially disturbed assembly of the complex as determined by sucrose gradient centrifugation and native PAGE. EPR spectroscopy of cytoplasmic membranes revealed that the BolA deletion results in the loss of the binuclear Fe/S cluster N1b.},
  language  = {en}
}
@article{TanabeLeimkuehlerDahl2019,
  author    = {Tanabe, Tomohisa Sebastian and Leimk{\"u}hler, Silke and Dahl, Christiane},
  title     = {The functional diversity of the prokaryotic sulfur carrier protein TusA},
  series = {Advances in microbial physiology},
  volume    = {75},
  journal   = {Advances in microbial physiology},
  editor    = {Poole, RK},
  publisher = {Elsevier Acad. Press},
  address   = {Amsterdam},
  isbn      = {978-0-12-817715-0},
  issn      = {0065-2911},
  doi       = {10.1016/bs.ampbs.2019.07.004},
  pages     = {233 -- 277},
  year      = {2019},
  abstract  = {Persulfide groups participate in a wide array of biochemical pathways and are chemically very versatile. The TusA protein has been identified as a central element supplying and transferring sulfur as persulfide to a number of important biosynthetic pathways, like molybdenum cofactor biosynthesis or thiomodifications in nucleosides of tRNAs. In recent years, it has furthermore become obvious that this protein is indispensable for the oxidation of sulfur compounds in the cytoplasm. Phylogenetic analyses revealed that different TusA protein variants exists in certain organisms, that have evolved to pursue specific roles in cellular pathways. The specific TusA-like proteins thereby cannot replace each other in their specific roles and are rather specific to one sulfur transfer pathway or shared between two pathways. While certain bacteria like Escherichia coli contain several copies of TusA-like proteins, in other bacteria like Allochromatium vinosum a single copy of TusA is present with an essential role for this organism. Here, we give an overview on the multiple roles of the various TusA-like proteins in sulfur transfer pathways in different organisms to shed light on the remaining mysteries of this versatile protein.},
  language  = {en}
}
@inproceedings{DuffusHartmannTeutloffetal.2019,
  author    = {Duffus, Benjamin R. and Hartmann, Tobias and Teutloff, Christian and Leimk{\"u}hler, Silke},
  title     = {Refining catalytic insights toward the chemical mechanism of R. capsulatus formate dehydrogenase via EPR spectroscopy},
  series = {Abstracts of papers : joint conference / The Chemical Institute of Cananda, CIC, American Chemical Society, ACS},
  volume    = {257},
  booktitle = {Abstracts of papers : joint conference / The Chemical Institute of Cananda, CIC, American Chemical Society, ACS},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0065-7727},
  pages     = {1},
  year      = {2019},
  language  = {en}
}
@article{MitrovaTadjoungWaffoKaufmannetal.2018,
  author    = {Mitrova, Biljana and Tadjoung Waffo, Armel Franklin and Kaufmann, Paul and Iobbi-Nivol, Chantal and Leimk{\"u}hler, Silke and Wollenberger, Ulla},
  title     = {Trimethylamine N-Oxide Electrochemical Biosensor with a Chimeric Enzyme},
  series = {ChemElectroChem},
  volume    = {6},
  journal   = {ChemElectroChem},
  number    = {6},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {2196-0216},
  doi       = {10.1002/celc.201801422},
  pages     = {1732 -- 1737},
  year      = {2018},
  abstract  = {For the first time, an enzyme-based electrochemical biosensor system for determination of trimethylamine N-oxide (TMAO) is described. It employs an active chimeric variant of TorA in combination with an enzymatically deoxygenating system and a low-potential mediator for effective regeneration of the enzyme and cathodic current generation. TMAO reductase (TorA) is a molybdoenzyme found in marine and most enterobacteria that specifically catalyzes the reduction of TMAO to trimethylamine (TMA). The chimeric TorA, named TorA-FDH, corresponds to the apoform of TorA from Escherichia coli reconstituted with the molybdenum cofactor from formate dehydrogenase (FDH). Each enzyme, TorA and TorA-FDH, was immobilized on the surface of a carbon electrode and protected with a dialysis membrane. The biosensor operates at an applied potential of -0.8V [vs. Ag/AgCl (1M KCl)] under ambient air conditions thanks to an additional enzymatic O-2-scavenger system. A comparison between the two enzymatic sensors revealed a much higher sensitivity for the biosensor with immobilized TorA-FDH. This biosensor exhibits a sensitivity of 14.16nA/M TMAO in a useful measuring range of 2-110M with a detection limit of LOD=2.96nM (S/N=3), and was similar for TMAO in buffer and in spiked serum samples. With a response time of 16 +/- 2 s, the biosensor is stable over prolonged daily measurements (n=20). This electrochemical biosensor provides suitable applications in detecting TMAO levels in human serum.},
  language  = {en}
}
@article{NeukranzKotterBeilschmidtetal.2019,
  author    = {Neukranz, Yannika and Kotter, Annika and Beilschmidt, Lena and Marelja, Zvonimir and Helm, Mark and Graf, Ralph and Leimk{\"u}hler, Silke},
  title     = {Analysis of the Cellular Roles of MOCS3 Identifies a MOCS3-Independent Localization of NFS1 at the Tips of the Centrosome},
  series = {Biochemistry},
  volume    = {58},
  journal   = {Biochemistry},
  number    = {13},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.8b01160},
  pages     = {1786 -- 1798},
  year      = {2019},
  abstract  = {The deficiency of the molybdenum cofactor (Moco) is an autosomal recessive disease, which leads to the loss of activity of all molybdoenzymes in humans with sulfite oxidase being the essential protein. Moco deficiency generally results in death in early childhood. Moco is a sulfur-containing cofactor synthesized in the cytosol with the sulfur being provided by a sulfur relay system composed of the L-cysteine desulfurase NFS1, MOCS3, and MOCS2A. Human MOCS3 is a dual-function protein that was shown to play an important role in Moco biosynthesis and in the mcm(5)s(2) U thio modifications of nucleosides in cytosolic tRNAs for Lys, Gln, and Glu. In this study, we constructed a homozygous MOCS3 knockout in HEK293T cells using the CRISPR/Cas9 system. The effects caused by the absence of MOCS3 were analyzed in detail. We show that sulfite oxidase activity was almost completely abolished, on the basis of the absence of Moco in these cells. In addition, mcm(5)s(2)U thio-modified tRNAs were not detectable. Because the L-cysteine desulfurase NFS1 was shown to act as a sulfur donor for MOCS3 in the cytosol, we additionally investigated the impact of a MOCS3 knockout on the cellular localization of NFS1. By different methods, we identified a MOCS3-independent novel localization of NFS1 at the centrosome.},
  language  = {en}
}
@article{ReschkeDuffusSchrapersetal.2019,
  author    = {Reschke, Stefan and Duffus, Benjamin R. and Schrapers, Peer and Mebs, Stefan and Teutloff, Christian and Dau, Holger and Haumann, Michael and Leimk{\"u}hler, Silke},
  title     = {Identification of YdhV as the First Molybdoenzyme Binding a Bis-Mo-MPT Cofactor in Escherichia coli},
  series = {Biochemistry},
  volume    = {58},
  journal   = {Biochemistry},
  number    = {17},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.9b00078},
  pages     = {2228 -- 2242},
  year      = {2019},
  abstract  = {The oxidoreductase YdhV in Escherichia coli has been predicted to belong to the family of molybdenum/tungsten cofactor (Moco/Wco)-containing enzymes. In this study, we characterized the YdhV protein in detail, which shares amino acid sequence homology with a tungsten-containing benzoyl-CoA reductase binding the bis-W-MPT (for metal-binding pterin) cofactor. The cofactor was identified to be of a bis-Mo-MPT type with no guanine nucleotides present, which represents a form of Moco that has not been found previously in any molybdoenzyme. Our studies showed that YdhV has a preference for bis-Mo-MPT over bis-W-MPT to be inserted into the enzyme. In-depth characterization of YdhV by X-ray absorption and electron paramagnetic resonance spectroscopies revealed that the bis-Mo-MPT cofactor in YdhV is redox active. The bis-Mo-MPT and bis-W-MPT cofactors include metal centers that bind the four sulfurs from the two dithiolene groups in addition to a cysteine and likely a sulfido ligand. The unexpected presence of a bis-Mo-MPT cofactor opens an additional route for cofactor biosynthesis in E. coli and expands the canon of the structurally highly versatile molybdenum and tungsten cofactors.},
  language  = {en}
}
@article{MotaEsmaeeliMoghaddamTabalvandaniCoelhoetal.2019,
  author    = {Mota, Cristiano and Esmaeeli Moghaddam Tabalvandani, Mariam and Coelho, Catarina and Santos-Silva, Teresa and Wolff, Martin and Foti, Alessandro and Leimk{\"u}hler, Silke and Romao, Maria Joao},
  title     = {Human aldehyde oxidase (hAOX1)},
  series = {FEBS Open Bio},
  volume    = {9},
  journal   = {FEBS Open Bio},
  number    = {5},
  publisher = {Wiley},
  address   = {Hoboken},
  issn      = {2211-5463},
  doi       = {10.1002/2211-5463.12617},
  pages     = {925 -- 934},
  year      = {2019},
  abstract  = {Human aldehyde oxidase (hAOX1) is a molybdenum enzyme with high toxicological importance, but its physiological role is still unknown. hAOX1 metabolizes different classes of xenobiotics and is one of the main drug-metabolizing enzymes in the liver, along with cytochrome P450. hAOX1 oxidizes and inactivates a large number of drug molecules and has been responsible for the failure of several phase I clinical trials. The interindividual variability of drug-metabolizing enzymes caused by single nucleotide polymorphisms (SNPs) is highly relevant in pharmaceutical treatments. In this study, we present the crystal structure of the inactive variant G1269R, revealing the first structure of a molybdenum cofactor (Moco)-free form of hAOX1. These data allowed to model, for the first time, the flexible Gate 1 that controls access to the active site. Furthermore, we inspected the thermostability of wild-type hAOX1 and hAOX1 with various SNPs (L438V, R1231H, G1269R or S1271L) by CD spectroscopy and ThermoFAD, revealing that amino acid exchanges close to the Moco site can impact protein stability up to 10 degrees C. These results correlated with biochemical and structural data and enhance our understanding of hAOX1 and the effect of SNPs in the gene encoding this enzyme in the human population. EnzymesAldehyde oxidase (); xanthine dehydrogenase (); xanthine oxidase (). DatabasesStructural data are available in the Protein Data Bank under the accession number .},
  language  = {en}
}
@article{BadalyanDierichStibaetal.2014,
  author    = {Badalyan, Artavazd and Dierich, Marlen and Stiba, Konstanze and Schwuchow, Viola and Leimk{\"u}hler, Silke and Wollenberger, Ulla},
  title     = {Electrical wiring of the aldehyde oxidoreductase PaoABC with a polymer containing osmium redox centers},
  series = {Biosensors},
  volume    = {4},
  journal   = {Biosensors},
  number    = {4},
  publisher = {MDPI},
  address   = {Basel},
  doi       = {10.3390/bios4040403},
  pages     = {403 -- 421},
  year      = {2014},
  abstract  = {Biosensors for the detection of benzaldehyde and g-aminobutyric acid (GABA) are reported using aldehyde oxidoreductase PaoABC from Escherichia coli immobilized in a polymer containing bound low potential osmium redox complexes. The electrically connected enzyme already electrooxidizes benzaldehyde at potentials below -0.15 V (vs. Ag|AgCl, 1 M KCl). The pH-dependence of benzaldehyde oxidation can be strongly influenced by the ionic strength. The effect is similar with the soluble osmium redox complex and therefore indicates a clear electrostatic effect on the bioelectrocatalytic efficiency of PaoABC in the osmium containing redox polymer. At lower ionic strength, the pH-optimum is high and can be switched to low pH-values at high ionic strength. This offers biosensing at high and low pH-values. A "reagentless" biosensor has been formed with enzyme wired onto a screen-printed electrode in a flow cell device. The response time to addition of benzaldehyde is 30 s, and the measuring range is between 10-150 µM and the detection limit of 5 µM (signal to noise ratio 3:1) of benzaldehyde. The relative standard deviation in a series (n = 13) for 200 µM benzaldehyde is 1.9\%. For the biosensor, a response to succinic semialdehyde was also identified. Based on this response and the ability to work at high pH a biosensor for GABA is proposed by coimmobilizing GABA-aminotransferase (GABA-T) and PaoABC in the osmium containing redox polymer.},
  language  = {en}
}
@misc{MogaRobinsonLeimkuehler2019,
  author    = {Moga, A. and Robinson, T. and Leimk{\"u}hler, Silke},
  title     = {Towards reconstituting a biosynthetic pathway within compartmentalized GUVs},
  series = {European biophysics journal : with biophysics letters ; an international journal of biophysics},
  volume    = {48},
  journal   = {European biophysics journal : with biophysics letters ; an international journal of biophysics},
  publisher = {Springer},
  address   = {New York},
  issn      = {0175-7571},
  pages     = {S218 -- S218},
  year      = {2019},
  language  = {en}
}
@article{TangWerchmeisterPredaetal.2019,
  author    = {Tang, Jing and Werchmeister, Rebecka Maria Larsen and Preda, Loredana and Huang, Wei and Zheng, Zhiyong and Leimk{\"u}hler, Silke and Wollenberger, Ulla and Xiao, Xinxin and Engelbrekt, Christian and Ulstrup, Jens and Zhang, Jingdong},
  title     = {Three-dimensional sulfite oxidase bioanodes based on graphene functionalized carbon paper for sulfite/O-2 biofuel cells},
  series = {ACS catalysis},
  volume    = {9},
  journal   = {ACS catalysis},
  number    = {7},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {2155-5435},
  doi       = {10.1021/acscatal.9b01715},
  pages     = {6543 -- 6554},
  year      = {2019},
  abstract  = {We have developed a three-dimensional (3D) graphene electrode suitable for the immobilization of human sulfite oxidase (hSO), which catalyzes the electrochemical oxidation of sulfite via direct electron transfer (DET). The electrode is fabricated by drop-casting graphene-polyethylenimine (G-P) composites on carbon papers (CPs) precoated with graphene oxide (GO). The negatively charged hSO can be adsorbed electrostatically on the positively charged matrix (G-P) on CP electrodes coated with GO (CPG), with a proper orientation for accelerated DET. Notably, further electrochemical reduction of G-P on CPG electrodes leads to a 9-fold increase of the saturation catalytic current density (j(m)) for sulfite oxidation reaching 24.4 +/- 0.3 mu A to cm(-2), the highest value among reported DET-based hSO bioelectrodes. The increased electron transfer rate plays a dominating role in the enhancement of direct enzymatic current because of the improved electric contact of hSO with the electrode, The optimized hSO bioelectrode shows a significant catalytic rate (k(cat): 25.6 +/- 0.3 s(-1)) and efficiency (k(cat)/K-m: 0.231 +/- 0.003 s(-1) mu M-1) compared to the reported hSO bioelectrodes. The assembly of the hSO bioanode and a commercial platinum biocathode allows the construction of sulfite/O-2 enzymatic biofuel cells (EBFCs) with flowing fuels. The optimized EBFC displays an open-circuit voltage (OCV) of 0.64 +/- 0.01 V and a maximum power density of 61 +/- 6 mu W cm(-2) (122 +/- 12 mW m(-3)) at 30 degrees C, which exceeds the best reported value by more than 6 times.},
  language  = {en}
}
@misc{BadalyanDierichStibaetal.2014,
  author    = {Badalyan, Artavazd and Dierich, Marlen and Stiba, Konstanze and Schwuchow, Viola and Leimk{\"u}hler, Silke and Wollenberger, Ulla},
  title     = {Electrical wiring of the aldehyde oxidoreductase PaoABC with a polymer containing osmium redox centers},
  series = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  number    = {1082},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-47507},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-475070},
  pages     = {21},
  year      = {2014},
  abstract  = {Biosensors for the detection of benzaldehyde and g-aminobutyric acid (GABA) are reported using aldehyde oxidoreductase PaoABC from Escherichia coli immobilized in a polymer containing bound low potential osmium redox complexes. The electrically connected enzyme already electrooxidizes benzaldehyde at potentials below -0.15 V (vs. Ag|AgCl, 1 M KCl). The pH-dependence of benzaldehyde oxidation can be strongly influenced by the ionic strength. The effect is similar with the soluble osmium redox complex and therefore indicates a clear electrostatic effect on the bioelectrocatalytic efficiency of PaoABC in the osmium containing redox polymer. At lower ionic strength, the pH-optimum is high and can be switched to low pH-values at high ionic strength. This offers biosensing at high and low pH-values. A "reagentless" biosensor has been formed with enzyme wired onto a screen-printed electrode in a flow cell device. The response time to addition of benzaldehyde is 30 s, and the measuring range is between 10-150 µM and the detection limit of 5 µM (signal to noise ratio 3:1) of benzaldehyde. The relative standard deviation in a series (n = 13) for 200 µM benzaldehyde is 1.9\%. For the biosensor, a response to succinic semialdehyde was also identified. Based on this response and the ability to work at high pH a biosensor for GABA is proposed by coimmobilizing GABA-aminotransferase (GABA-T) and PaoABC in the osmium containing redox polymer.},
  language  = {en}
}
@article{ZupokGorkaSiemiatkowskaetal.2019,
  author    = {Zupok, Arkadiusz and G{\´o}rka, Michał Jakub and Siemiatkowska, Beata and Skirycz, Aleksandra and Leimk{\"u}hler, Silke},
  title     = {Iron-Dependent Regulation of Molybdenum Cofactor Biosynthesis Genes in Escherichia coli},
  series = {Journal of bacteriology},
  volume    = {201},
  journal   = {Journal of bacteriology},
  number    = {17},
  publisher = {American Society for Microbiology},
  address   = {Washington},
  issn      = {0021-9193},
  doi       = {10.1128/JB.00382-19},
  pages     = {15},
  year      = {2019},
  abstract  = {Molybdenum cofactor (Moco) biosynthesis is a complex process that involves the coordinated function of several proteins. In recent years it has become obvious that the availability of iron plays an important role in the biosynthesis of Moco. First, the MoaA protein binds two (4Fe-4S] clusters per monomer. Second, the expression of the moaABCDE and moeAB operons is regulated by FNR, which senses the availability of oxygen via a functional NFe-4S) cluster. Finally, the conversion of cyclic pyranopterin monophosphate to molybdopterin requires the availability of the L-cysteine desulfurase IscS, which is a shared protein with a main role in the assembly of Fe-S clusters. In this report, we investigated the transcriptional regulation of the moaABCDE operon by focusing on its dependence on cellular iron availability. While the abundance of selected molybdoenzymes is largely decreased under iron-limiting conditions, our data show that the regulation of the moaABCDE operon at the level of transcription is only marginally influenced by the availability of iron. Nevertheless, intracellular levels of Moco were decreased under iron-limiting conditions, likely based on an inactive MoaA protein in addition to lower levels of the L-cysteine desulfurase IscS, which simultaneously reduces the sulfur availability for Moco production. IMPORTANCE FNR is a very important transcriptional factor that represents the master switch for the expression of target genes in response to anaerobiosis. Among the FNR-regulated operons in Escherichia coli is the moaABCDE operon, involved in Moco biosynthesis. Molybdoenzymes have essential roles in eukaryotic and prokaryotic organisms. In bacteria, molybdoenzymes are crucial for anaerobic respiration using alternative electron acceptors. This work investigates the connection of iron availability to the biosynthesis of Moco and the production of active molybdoenzymes.},
  language  = {en}
}
@article{LemaireHonoreTempeletal.2019,
  author    = {Lemaire, Olivier N. and Honore, Flora A. and Tempel, Sebastien and Fortier, Emma M. and Leimk{\"u}hler, Silke and Mejean, Vincent and Iobbi-Nivol, Chantal},
  title     = {Shewanella decolorationis LDS1 Chromate Resistance},
  series = {Applied and environmental microbiology},
  volume    = {85},
  journal   = {Applied and environmental microbiology},
  number    = {18},
  publisher = {American Society for Microbiology},
  address   = {Washington},
  issn      = {0099-2240},
  doi       = {10.1128/AEM.00777-19},
  pages     = {15},
  year      = {2019},
  abstract  = {The genus Shewanella is well known for its genetic diversity, its outstanding respiratory capacity, and its high potential for bioremediation. Here, a novel strain isolated from sediments of the Indian Ocean was characterized. A 16S rRNA analysis indicated that it belongs to the species Shewanella decolorationis. It was named Shewanella decolorationis LDS1. This strain presented an unusual ability to grow efficiently at temperatures from 24 degrees C to 40 degrees C without apparent modifications of its metabolism, as shown by testing respiratory activities or carbon assimilation, and in a wide range of salt concentrations. Moreover, S. decolorationis LDS1 tolerates high chromate concentrations. Indeed, it was able to grow in the presence of 4 mM chromate at 28 degrees C and 3 mM chromate at 40 degrees C. Interestingly, whatever the temperature, when the culture reached the stationary phase, the strain reduced the chromate present in the growth medium. In addition, S. decolorationis LDS1 degrades different toxic dyes, including anthraquinone, triarylmethane, and azo dyes. Thus, compared to Shewanella oneidensis, this strain presented better capacity to cope with various abiotic stresses, particularly at high temperatures. The analysis of genome sequence preliminary data indicated that, in contrast to S. oneidensis and S. decolorationis S12, S. decolorationis LDS1 possesses the phosphorothioate modification machinery that has been described as participating in survival against various abiotic stresses by protecting DNA. We demonstrate that its heterologous production in S. oneidensis allows it to resist higher concentrations of chromate. IMPORTANCE Shewanella species have long been described as interesting microorganisms in regard to their ability to reduce many organic and inorganic compounds, including metals. However, members of the Shewanella genus are often depicted as cold-water microorganisms, although their optimal growth temperature usually ranges from 25 to 28 degrees C under laboratory growth conditions. Shewanella decolorationis LDS1 is highly attractive, since its metabolism allows it to develop efficiently at temperatures from 24 to 40 degrees C, conserving its ability to respire alternative substrates and to reduce toxic compounds such as chromate or toxic dyes. Our results clearly indicate that this novel strain has the potential to be a powerful tool for bioremediation and unveil one of the mechanisms involved in its chromate resistance.},
  language  = {en}
}
@misc{LeimkuehlerBuehningBeilschmidt2017,
  author    = {Leimk{\"u}hler, Silke and B{\"u}hning, Martin and Beilschmidt, Lena},
  title     = {Shared sulfur mobilization routes for tRNA thiolation and molybdenum cofactor biosynthesis in prokaryotes and eukaryotes},
  series = {Postprints der Universit{\"a}t Potsdam : Mathematisch Naturwissenschaftliche Reihe},
  journal   = {Postprints der Universit{\"a}t Potsdam : Mathematisch Naturwissenschaftliche Reihe},
  number    = {1015},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-47501},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-475011},
  pages     = {22},
  year      = {2017},
  abstract  = {Modifications of transfer RNA (tRNA) have been shown to play critical roles in the biogenesis, metabolism, structural stability and function of RNA molecules, and the specific modifications of nucleobases with sulfur atoms in tRNA are present in pro- and eukaryotes. Here, especially the thiomodifications xm(5)s(2)U at the wobble position 34 in tRNAs for Lys, Gln and Glu, were suggested to have an important role during the translation process by ensuring accurate deciphering of the genetic code and by stabilization of the tRNA structure. The trafficking and delivery of sulfur nucleosides is a complex process carried out by sulfur relay systems involving numerous proteins, which not only deliver sulfur to the specific tRNAs but also to other sulfur-containing molecules including iron-sulfur clusters, thiamin, biotin, lipoic acid and molybdopterin (MPT). Among the biosynthesis of these sulfur-containing molecules, the biosynthesis of the molybdenum cofactor (Moco) and the synthesis of thio-modified tRNAs in particular show a surprising link by sharing protein components for sulfur mobilization in pro- and eukaryotes.},
  language  = {en}
}
@misc{ZupokIobbiNivolMejeanetal.2019,
  author    = {Zupok, Arkadiusz and Iobbi-Nivol, Chantal and Mejean, Vincent and Leimk{\"u}hler, Silke},
  title     = {The regulation of Moco biosynthesis and molybdoenzyme gene expression by molybdenum and iron in bacteria},
  series = {Metallomics : integrated biometal science},
  volume    = {11},
  journal   = {Metallomics : integrated biometal science},
  number    = {10},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  issn      = {1756-5901},
  doi       = {10.1039/c9mt00186g},
  pages     = {1602 -- 1624},
  year      = {2019},
  abstract  = {Bacterial molybdoenzymes are key enzymes involved in the global sulphur, nitrogen and carbon cycles. These enzymes require the insertion of the molybdenum cofactor (Moco) into their active sites and are able to catalyse a large range of redox-reactions. Escherichia coli harbours nineteen different molybdoenzymes that require a tight regulation of their synthesis according to substrate availability, oxygen availability and the cellular concentration of molybdenum and iron. The synthesis and assembly of active molybdoenzymes are regulated at the level of transcription of the structural genes and of translation in addition to the genes involved in Moco biosynthesis. The action of global transcriptional regulators like FNR, NarXL/QP, Fur and ArcA and their roles on the expression of these genes is described in detail. In this review we focus on what is known about the molybdenum- and iron-dependent regulation of molybdoenzyme and Moco biosynthesis genes in the model organism E. coli. The gene regulation in E. coli is compared to two other well studied model organisms Rhodobacter capsulatus and Shewanella oneidensis.},
  language  = {en}
}
@misc{OttoMareljaSchoofsetal.2018,
  author    = {Otto, Nils and Marelja, Zvonimir and Schoofs, Andreas and Kranenburg, Holger and Bittern, Jonas and Yildirim, Kerem and Berh, Dimitri and Bethke, Maria and Thomas, Silke and Rode, Sandra and Risse, Benjamin and Jiang, Xiaoyi and Pankratz, Michael and Leimk{\"u}hler, Silke and Kl{\"a}mbt, Christian},
  title     = {The sulfite oxidase Shopper controls neuronal activity by regulating glutamate homeostasis in Drosophila ensheathing glia},
  series = {Postprints der Universit{\"a}t Potsdam : Mathematisch Naturwissenschaftliche Reihe},
  journal   = {Postprints der Universit{\"a}t Potsdam : Mathematisch Naturwissenschaftliche Reihe},
  number    = {975},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-42620},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-426205},
  pages     = {14},
  year      = {2018},
  abstract  = {Specialized glial subtypes provide support to developing and functioning neural networks. Astrocytes modulate information processing by neurotransmitter recycling and release of neuromodulatory substances, whereas ensheathing glial cells have not been associated with neuromodulatory functions yet. To decipher a possible role of ensheathing glia in neuronal information processing, we screened for glial genes required in the Drosophila central nervous system for normal locomotor behavior. Shopper encodes a mitochondrial sulfite oxidase that is specifically required in ensheathing glia to regulate head bending and peristalsis. shopper mutants show elevated sulfite levels affecting the glutamate homeostasis which then act on neuronal network function. Interestingly, human patients lacking the Shopper homolog SUOX develop neurological symptoms, including seizures. Given an enhanced expression of SUOX by oligodendrocytes, our findings might indicate that in both invertebrates and vertebrates more than one glial cell type may be involved in modulating neuronal activity.},
  language  = {en}
}
@article{HahnewaldLeimkuehlerVilasecaetal.2006,
  author    = {Hahnewald, Rita and Leimk{\"u}hler, Silke and Vilaseca, Antonia and Acquaviva-Bourdain, Cecile and Lenz, Ulrike and Reiss, Jochen},
  title     = {A novel MOCS2 mutation reveals coordinated expression of the small and large subunit of molybdopterin synthase},
  series = {Molecular genetics and metabolism},
  volume    = {89},
  journal   = {Molecular genetics and metabolism},
  number    = {3},
  publisher = {Elsevier},
  address   = {San Diego},
  issn      = {1096-7192},
  doi       = {10.1016/j.ymgme.2006.04.008},
  pages     = {210 -- 213},
  year      = {2006},
  abstract  = {The small and large subunits of molybdopterin (MPT) synthase (MOCS2A and MOCS2B), are both encoded by the MOCS2 gene in overlapping and shifted open reading frames (ORFs), which is a highly unusual structure for eukaryotes. Theoretical analysis of genomic sequences suggested that the expression of these overlapping ORFs is facilitated by the use of alternate first exons leading to alternative transcripts. Here, we confirm the existence of these overlapping transcripts experimentally. Further, we identified a deletion in a molybdenum cofactor deficient patient, which removes the start codon for the small subunit (MOCS2A). We observed undisturbed production of both transcripts, while Western blot analysis demonstrated that MOCS2B, the large subunit, is unstable in the absence of MOCS2A. This reveals new insights into the expression of this evolutionary ancient anabolic system.},
  language  = {en}
}
@article{DongYangReschkeetal.2017,
  author    = {Dong, Chao and Yang, Jing and Reschke, Stefan and Leimk{\"u}hler, Silke and Kirk, Martin L.},
  title     = {Vibrational Probes of Molybdenum Cofactor-Protein Interactions in Xanthine Dehydrogenase},
  series = {Inorganic chemistry},
  volume    = {56},
  journal   = {Inorganic chemistry},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0020-1669},
  doi       = {10.1021/acs.inorgchem.7b00028},
  pages     = {6830 -- 6837},
  year      = {2017},
  abstract  = {The pyranopterin dithiolene (PDT) ligand is an integral component of the molybdenum cofactor (Moco) found in all molybdoenzymes with the sole exception of nitrogenase. However, the roles of the PDT in catalysis are still unknown. The PDT is believed to be bound to the proteins by an extensive hydrogen bonding network, and it has been suggested that these interactions may function to fine-tune Moco for electron- and atom-transfer reactivity in catalysis. Here, we use resonance Raman (rR) spectroscopy to probe Moco-protein interactions using heavy-atom congeners of lumazine, molecules that bind tightly to both wild-type xanthine dehydrogenase (wt-XDH) and its Q102G and Q197A variants following enzymatic hydroxylation to the corresponding violapterin product molecules. The resulting enzyme-product complexes possess intense near-IR absorption, allowing high-quality rR spectra to be collected on wt-XDH and the Q102G and Q197A variants. Small negative frequency shifts relative to wt-XDH are observed for the low-frequency Moco vibrations. These results are interpreted in the context of weak hydrogen-bonding and/or electrostatic interactions between Q102 and the -NH2 terminus of the PDT, and between Q197 and the terminal oxo of the Mo equivalent to O group. The Q102A, Q102G, Q197A, and Q197E variants do not appreciably affect the kinetic parameters k(red) and k(red)/K-D, indicating that a primary role for these glutamine residues is to stabilize and coordinate Moco in the active site of XO family enzymes but to not directly affect the catalytic throughput. Raman frequency shifts between wt-XDH and its Q102G variant suggest that the changes in the electron density at the Mo ion that accompany Mo oxidation during electron-transfer regeneration of the catalytically competent active site are manifest in distortions at the distant PDT amino terminus. This implies a primary role for the PDT as a conduit for facilitating enzymatic electron-transfer reactivity in xanthine oxidase family enzymes.},
  language  = {en}
}
@article{FotiDorendorfLeimkuehler2017,
  author    = {Foti, Alessandro and Dorendorf, Frank and Leimk{\"u}hler, Silke},
  title     = {A single nucleotide polymorphism causes enhanced radical oxygen species production by human aldehyde oxidase},
  series = {PLoS one},
  volume    = {12},
  journal   = {PLoS one},
  publisher = {PLoS},
  address   = {San Fransisco},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0182061},
  pages     = {18338 -- 18347},
  year      = {2017},
  abstract  = {Aldehyde oxidases (AOXs) are molybdo-flavoenzymes characterized by broad substrate specificity, oxidizing aromatic/aliphatic aldehydes into the corresponding carboxylic acids and hydroxylating various heteroaromatic rings. The enzymes use oxygen as the terminal electron acceptor and produce reduced oxygen species during turnover. The physiological function of mammalian AOX isoenzymes is still unclear, however, human AOX (hAOX1) is an emerging enzyme in phase-I drug metabolism. Indeed, the number of xenobiotics acting as hAOX1 substrates is increasing. Further, numerous single-nucleotide polymorphisms (SNPs) have been identified within the hAOX1 gene. SNPs are a major source of inter-individual variability in the human population, and SNP-based amino acid exchanges in hAOX1 reportedly modulate the catalytic function of the enzyme in either a positive or negative fashion. In this report we selected ten novel SNPs resulting in amino acid exchanges in proximity to the FAD site of hAOX1 and characterized the purified enzymes after heterologous expression in Escherichia coli. The hAOX1 variants were characterized carefully by quantitative differences in their ability to produce superoxide radical. ROS represent prominent key molecules in physiological and pathological conditions in the cell. Our data reveal significant alterations in superoxide anion production among the variants. In particular the SNP-based amino acid exchange L438V in proximity to the isoalloxanzine ring of the FAD cofactor resulted in increased rate of superoxide radical production of 75\%. Considering the high toxicity of the superoxide in the cell, the hAOX1-L438V SNP variant is an eventual candidate for critical or pathological roles of this natural variant within the human population.},
  language  = {en}
}
@article{KuecuekgoezeTeraoGarattinietal.2017,
  author    = {Kuecuekgoeze, Goekhan and Terao, Mineko and Garattini, Enrico and Leimk{\"u}hler, Silke},
  title     = {Direct Comparison of the Enzymatic Characteristics and Superoxide Production of the Four Aldehyde Oxidase Enzymes Present in Mouse},
  series = {Drug metabolism and disposition : the biological fate of chemicals},
  volume    = {45},
  journal   = {Drug metabolism and disposition : the biological fate of chemicals},
  publisher = {American Society for Pharmacology and Experimental Therapeutics},
  address   = {Bethesda},
  issn      = {0090-9556},
  doi       = {10.1124/dmd.117.075937},
  pages     = {947 -- 955},
  year      = {2017},
  abstract  = {Aldehyde oxidases (AOXs) are molybdoflavoenzymes with an important role in the metabolism and detoxification of heterocyclic compounds and aliphatic as well as aromatic aldehydes. The enzymes use oxygen as the terminal electron acceptor and produce reduced oxygen species during turnover. Four different enzymes, mAOX1, mAOX3, mAOX4, and mAOX2, which are the products of distinct genes, are present in the mouse. A direct and simultaneous comparison of the enzymatic properties and characteristics of the four enzymes has never been performed. In this report, the four catalytically active mAOX enzymes were purified after heterologous expression in Escherichia coli. The kinetic parameters of the four mouse AOX enzymes were determined and compared with the use of six predicted substrates of physiologic and toxicological interest, i.e., retinaldehyde, N1-methylnicotinamide, pyridoxal, vanillin, 4-(dimethylamino) cinnamaldehyde (p-DMAC), and salicylaldehyde. While retinaldehyde, vanillin, p-DMAC, and salycilaldehyde are efficient substrates for the four mouse AOX enzymes, N1-methylnicotinamide is not a substrate of mAOX1 or mAOX4, and pyridoxal is notmetabolized by any of the purified enzymes. Overall, mAOX1, mAOX2, mAOX3, and mAOX4 are characterized by significantly different KM and kcat values for the active substrates. The four mouse AOXs are also characterized by quantitative differences in their ability to produce superoxide radicals. With respect to this last point, mAOX2 is the enzyme generating the largest rate of superoxide radicals of around 40\% in relation to moles of substrate converted, and mAOX1, the homolog to the human enzyme, produces a rate of approximately 30\% of superoxide radicals with the same substrate.},
  language  = {en}
}
@article{ZengLeimkuehlerWollenbergeretal.2017,
  author    = {Zeng, Ting and Leimk{\"u}hler, Silke and Wollenberger, Ulla and Fourmond, Vincent},
  title     = {Transient Catalytic Voltammetry of Sulfite Oxidase Reveals Rate Limiting Conformational Changes},
  series = {Journal of the American Chemical Society},
  volume    = {139},
  journal   = {Journal of the American Chemical Society},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0002-7863},
  doi       = {10.1021/jacs.7b05480},
  pages     = {11559 -- 11567},
  year      = {2017},
  abstract  = {Sulfite oxidases are metalloenzymes that oxidize sulfite to sulfate at a molybdenum active site. In vertebrate sulfite oxidases, the electrons generated at the Mo center are transferred to an external electron acceptor via a heme domain, which can adopt two conformations: a "closed" conformation, suitable for internal electron transfer, and an "open" conformation suitable for intermolecular electron transfer. This conformational change is an integral part of the catalytic cycle. Sulfite oxidases have been wired to electrode surfaces, but their immobilization leads to a significant decrease in their catalytic activity, raising the question of the occurrence of the conformational change when the enzyme is on an electrode. We recorded and quantitatively modeled for the first time the transient response of the catalytic cycle of human sulfite oxidase immobilized on an electrode. We show that conformational changes still occur on the electrode, but at a lower rate than in solution, which is the reason for the decrease in activity of sulfite oxidases upon immobilization.},
  language  = {en}
}
@article{BuehningFriemelLeimkuehler2017,
  author    = {B{\"u}hning, Martin and Friemel, Martin and Leimk{\"u}hler, Silke},
  title     = {Functional Complementation Studies Reveal Different Interaction Partners of Escherichia coil IscS and Human NFS1},
  series = {Biochemistry},
  volume    = {56},
  journal   = {Biochemistry},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.7b00627},
  pages     = {4592 -- 4605},
  year      = {2017},
  abstract  = {The trafficking and delivery of sulfur to cofactors and nucleosides is a highly regulated and conserved process among all organisms. All sulfur transfer pathways generally have an L-cysteine desulfurase as an initial sulfur mobilizing enzyme in common, which serves as a sulfur donor for the biosynthesis of sulfur-containing biomolecules like iron sulfur (Fe-S) clusters, thiamine, biotin, lipoic acid, the molybdenum cofactor (Moco), and thiolated nucleosides in tRNA. The human L-cysteine desulfurase NFS1 and the Escherichia coli homologue IscS share a level of amino acid sequence identity of similar to 60\%. While E. coli IscS has a versatile role in the cell and was shown to have numerous interaction partners, NFS1 is mainly localized in mitochondria with a crucial role in the biosynthesis of Fe-S clusters. Additionally, NFS1 is also located in smaller amounts in the cytosol with a role in Moco biosynthesis and mcm(5)s(2)U34 thio modifications of nucleosides in tRNA. NFS1 and IscS were conclusively shown to have different interaction partners in their respective organisms. Here, we used functional complementation studies of an E. coli iscS deletion strain with human NFS1 to dissect their conserved roles in the transfer of sulfur to a specific target protein. Our results show that human NFS1 and E. coli IscS share conserved binding sites for proteins involved in Fe-S cluster assembly like IscU, but not with proteins for tRNA thio modifications or Moco biosynthesis. In addition, we show that human NFS1 was almost fully able to complement the role of IscS in Moco biosynthesis when its specific interaction partner protein MOCS3 from humans was also present.},
  language  = {en}
}
@misc{Leimkuehler2017,
  author    = {Leimk{\"u}hler, Silke},
  title     = {Shared function and moonlighting proteins in molybdenum cofactor biosynthesis},
  series = {Biological chemistry},
  volume    = {398},
  journal   = {Biological chemistry},
  publisher = {De Gruyter},
  address   = {Berlin},
  issn      = {1431-6730},
  doi       = {10.1515/hsz-2017-0110},
  pages     = {1009 -- 1026},
  year      = {2017},
  abstract  = {The biosynthesis of the molybdenum cofactor (Moco) is a highly conserved pathway in bacteria, archaea and eukaryotes. The molybdenum atom in Moco-containing enzymes is coordinated to the dithiolene group of a tricyclic pyranopterin monophosphate cofactor. The biosynthesis of Moco can be divided into three conserved steps, with a fourth present only in bacteria and archaea: (1) formation of cyclic pyranopterin monophosphate, (2) formation of molybdopterin (MPT), (3) insertion of molybdenum into MPT to form Mo-MPT, and (4) additional modification of Mo-MPT in bacteria with the attachment of a GMP or CMP nucleotide, forming the dinucleotide variants of Moco. While the proteins involved in the catalytic reaction of each step of Moco biosynthesis are highly conserved among the Phyla, a surprising link to other cellular pathways has been identified by recent discoveries. In particular, the pathways for FeS cluster assembly and thio-modifications of tRNA are connected to Moco biosynthesis by sharing the same protein components. Further, proteins involved in Moco biosynthesis are not only shared with other pathways, but additionally have moonlighting roles. This review gives an overview of Moco biosynthesis in bacteria and humans and highlights the shared function and moonlighting roles of the participating proteins.},
  language  = {en}
}
@article{BrietzkeDietzKellingetal.2017,
  author    = {Brietzke, Thomas Martin and Dietz, Thomas and Kelling, Alexandra and Schilde, Uwe and Bois, Juliana and Kelm, Harald and Reh, Manuel and Schmitz, Markus and Koerzdoerfer, Thomas and Leimk{\"u}hler, Silke and Wollenberger, Ulla and Krueger, Hans-Joerg and Holdt, Hans-J{\"u}rgen},
  title     = {The 1,6,7,12-Tetraazaperylene Bridging Ligand as an Electron Reservoir and Its Disulfonato Derivative as Redox Mediator in an Enzyme-Electrode Process},
  series = {Chemistry - a European journal},
  volume    = {23},
  journal   = {Chemistry - a European journal},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {0947-6539},
  doi       = {10.1002/chem.201703639},
  pages     = {15583 -- 15587},
  year      = {2017},
  abstract  = {The homodinuclear ruthenium(II) complex [{Ru(l-N4Me2)}(2)(-tape)](PF6)(4) {[1](PF6)(4)} (l-N4Me2=N,N-dimethyl-2,11-diaza[3.3](2,6)-pyridinophane, tape=1,6,7,12-tetraazaperylene) can store one or two electrons in the energetically low-lying * orbital of the bridging ligand tape. The corresponding singly and doubly reduced complexes [{Ru(l-N4Me2)}(2)(-tape(.-))](PF6)(3) {[2](PF6)(3)} and [{Ru(l-N4Me2)}(2)(-tape(2-))](PF6)(2) {[3](PF6)(2)}, respectively, were electrochemically generated, successfully isolated and fully characterized by single-crystal X-ray crystallography, spectroscopic methods and magnetic susceptibility measurements. The singly reduced complex [2](PF6)(3) contains the -radical tape(.-) and the doubly reduced [3](PF6)(2) the diamagnetic dianion tape(2-) as bridging ligand, respectively. Nucleophilic aromatic substitution at the bridging tape in [1](4+) by two sulfite units gave the complex [{Ru(l-N4Me2)}(2){-tape-(SO3)(2)}](2+) ([4](2+)). Complex dication [4](2+) was exploited as a redox mediator between an anaerobic homogenous reaction solution of an enzyme system (sulfite/sulfite oxidase) and the electrode via participation of the low-energy *-orbital of the disulfonato-substituted bridging ligand tape-(SO3)(2)(2-) (E-red1=-0.1V versus Ag/AgCl/1m KCl in water).},
  language  = {en}
}
@misc{LeimkuehlerIobbiNivol2016,
  author    = {Leimk{\"u}hler, Silke and Iobbi-Nivol, Chantal},
  title     = {Bacterial molybdoenzymes: old enzymes for new purposes},
  series = {FEMS microbiology reviews},
  volume    = {40},
  journal   = {FEMS microbiology reviews},
  publisher = {Oxford Univ. Press},
  address   = {Oxford},
  issn      = {0168-6445},
  doi       = {10.1093/femsre/fuv043},
  pages     = {1 -- 18},
  year      = {2016},
  abstract  = {Molybdoenzymes are widespread in eukaryotic and prokaryotic organisms where they play crucial functions in detoxification reactions in the metabolism of humans and bacteria, in nitrate assimilation in plants and in anaerobic respiration in bacteria. To be fully active, these enzymes require complex molybdenum-containing cofactors, which are inserted into the apoenzymes after folding. For almost all the bacterial molybdoenzymes, molybdenum cofactor insertion requires the involvement of specific chaperones. In this review, an overview on the molybdenum cofactor biosynthetic pathway is given together with the role of specific chaperones dedicated for molybdenum cofactor insertion and maturation. Many bacteria are involved in geochemical cycles on earth and therefore have an environmental impact. The roles of molybdoenzymes in bioremediation and for environmental applications are presented.This review gives an overview of the diverse mechanisms leading to the insertion of the different forms of the molybdenum cofactor into the respective target enzymes and summarizes the roles of different molybdoenzymes in the environment.This review gives an overview of the diverse mechanisms leading to the insertion of the different forms of the molybdenum cofactor into the respective target enzymes and summarizes the roles of different molybdoenzymes in the environment.},
  language  = {en}
}
@misc{TeraoRomaoLeimkuehleretal.2016,
  author    = {Terao, Mineko and Romao, Maria Joao and Leimk{\"u}hler, Silke and Bolis, Marco and Fratelli, Maddalena and Coelho, Catarina and Santos-Silva, Teresa and Garattini, Enrico},
  title     = {Structure and function of mammalian aldehyde oxidases},
  series = {Archives of toxicology : official journal of EUROTOX},
  volume    = {90},
  journal   = {Archives of toxicology : official journal of EUROTOX},
  publisher = {Springer},
  address   = {Heidelberg},
  issn      = {0340-5761},
  doi       = {10.1007/s00204-016-1683-1},
  pages     = {753 -- 780},
  year      = {2016},
  abstract  = {Mammalian aldehyde oxidases (AOXs; EC1.2.3.1) are a group of conserved proteins belonging to the family of molybdo-flavoenzymes along with the structurally related xanthine dehydrogenase enzyme. AOXs are characterized by broad substrate specificity, oxidizing not only aromatic and aliphatic aldehydes into the corresponding carboxylic acids, but also hydroxylating a series of heteroaromatic rings. The number of AOX isoenzymes expressed in different vertebrate species is variable. The two extremes are represented by humans, which express a single enzyme (AOX1) in many organs and mice or rats which are characterized by tissue-specific expression of four isoforms (AOX1, AOX2, AOX3, and AOX4). In vertebrates each AOX isoenzyme is the product of a distinct gene consisting of 35 highly conserved exons. The extant species-specific complement of AOX isoenzymes is the result of a complex evolutionary process consisting of a first phase characterized by a series of asynchronous gene duplications and a second phase where the pseudogenization and gene deletion events prevail. In the last few years remarkable advances in the elucidation of the structural characteristics and the catalytic mechanisms of mammalian AOXs have been made thanks to the successful crystallization of human AOX1 and mouse AOX3. Much less is known about the physiological function and physiological substrates of human AOX1 and other mammalian AOX isoenzymes, although the importance of these proteins in xenobiotic metabolism is fairly well established and their relevance in drug development is increasing. This review article provides an overview and a discussion of the current knowledge on mammalian AOX.},
  language  = {en}
}
@article{HartmannSchrapersUteschetal.2016,
  author    = {Hartmann, Tobias and Schrapers, Peer and Utesch, Tillmann and Nimtz, Manfred and Rippers, Yvonne and Dau, Holger and Mroginski, Maria Andrea and Haumann, Michael and Leimk{\"u}hler, Silke},
  title     = {The Molybdenum Active Site of Formate Dehydrogenase Is Capable of Catalyzing C-H Bond Cleavage and Oxygen Atom Transfer Reactions},
  series = {Biochemistry},
  volume    = {55},
  journal   = {Biochemistry},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0006-2960},
  doi       = {10.1021/acs.biochem.6b00002},
  pages     = {2381 -- 2389},
  year      = {2016},
  abstract  = {Formate dehydrogenases (FDHs) are capable of performing the reversible oxidation of formate and are enzymes of great interest for fuel cell applications and for the production of reduced carbon compounds as energy sources from CO2. Metal containing FDHs in general contain a highly conserved active site, comprising a molybdenum (or tungsten) center coordinated by two molybdopterin guanine dinucleotide molecules, a sulfido and a (seleno-)cysteine ligand, in addition to a histidine and arginine residue in the second coordination sphere. So far, the role of these amino acids in catalysis has not been studied in detail, because of the lack of suitable expression systems and the lability or oxygen sensitivity of the enzymes. Here, the roles of these active site residues is revealed using the Mo-containing FDH from Rhodobacter capsulatus. Our results show that the cysteine ligand at the Mo ion is displaced by the formate substrate during the reaction, the arginine has a direct role in substrate binding and stabilization, and the histidine elevates the pK(a) of the active site cysteine. We further found that in addition to reversible formate oxidation, the enzyme is further capable of reducing nitrate to nitrite. We propose a mechanistic scheme that combines both functionalities and provides important insights into the distinct mechanisms of C-H bond cleavage and oxygen atom transfer catalyzed by formate dehydrogenase.},
  language  = {en}
}
@article{PinyouRuffPoelleretal.2016,
  author    = {Pinyou, Piyanut and Ruff, Adrian and Poeller, Sascha and Alsaoub, Sabine and Leimk{\"u}hler, Silke and Wollenberger, Ursula and Schuhmann, Wolfgang},
  title     = {Wiring of the aldehyde oxidoreductase PaoABC to electrode surfaces via entrapment in low potential phenothiazine-modified redox polymers},
  series = {Bioelectrochemistry : an international journal devoted to electrochemical aspects of biology and biological aspects of electrochemistry ; official journal of the Bioelectrochemical Society},
  volume    = {109},
  journal   = {Bioelectrochemistry : an international journal devoted to electrochemical aspects of biology and biological aspects of electrochemistry ; official journal of the Bioelectrochemical Society},
  publisher = {Elsevier},
  address   = {Lausanne},
  issn      = {1567-5394},
  doi       = {10.1016/j.bioelechem.2015.12.005},
  pages     = {24 -- 30},
  year      = {2016},
  abstract  = {Phenothiazine-modified redox hydrogels were synthesized and used for the wiring of the aldehyde oxidoreductase PaoABC to electrode surfaces. The effects of the pH value and electrode surface modification on the biocatalytic activity of the layers were studied in the presence of vanillin as the substrate. The enzyme electrodes were successfully employed as bioanodes in vanillin/O-2 biofuel cells in combination with a high potential bilirubin oxidase biocathode. Open circuit voltages of around 700 mV could be obtained in a two compartment biofuel cell setup. Moreover, the use of a rather hydrophobic polymer with a high degree of crosslinking sites ensures the formation of stable polymer/enzyme films which were successfully used as bioanode in membrane-less biofuel cells. (C) 2015 Elsevier B.V. All rights reserved.},
  language  = {en}
}
@article{YanFriemelAloisietal.2016,
  author    = {Yan, Robert and Friemel, Martin and Aloisi, Claudia and Huynen, Martijn and Taylor, Ian A. and Leimk{\"u}hler, Silke and Pastore, Annalisa},
  title     = {The Eukaryotic-Specific ISD11 Is a Complex-Orphan Protein with Ability to Bind the Prokaryotic IscS},
  series = {PLoS one},
  volume    = {11},
  journal   = {PLoS one},
  publisher = {PLoS},
  address   = {San Fransisco},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0157895},
  pages     = {383 -- 395},
  year      = {2016},
  abstract  = {The eukaryotic protein Isd11 is a chaperone that binds and stabilizes the central component of the essential metabolic pathway responsible for formation of iron-sulfur clusters in mitochondria, the desulfurase Nfs1. Little is known about the exact role of Isd11. Here, we show that human Isd11 (ISD11) is a helical protein which exists in solution as an equilibrium between monomer, dimeric and tetrameric species when in the absence of human Nfs1 (NFS1). We also show that, surprisingly, recombinant ISD11 expressed in E. coli co-purifies with the bacterial orthologue of NFS1, IscS. Binding is weak but specific suggesting that, despite the absence of Isd11 sequences in bacteria, there is enough conservation between the two desulfurases to retain a similar mode of interaction. This knowledge may inform us on the conservation of the mode of binding of Isd11 to the desulfurase. We used evolutionary evidence to suggest Isd11 residues involved in the interaction.},
  language  = {en}
}
@article{FotiHartmannCoelhoetal.2016,
  author    = {Foti, Alessandro and Hartmann, Tobias and Coelho, Catarina and Santos-Silva, Teresa and Romao, Maria Joao and Leimk{\"u}hler, Silke},
  title     = {Optimization of the Expression of Human Aldehyde Oxidase for Investigations of Single-Nucleotide Polymorphisms},
  series = {Drug metabolism and disposition : the biological fate of chemicals},
  volume    = {44},
  journal   = {Drug metabolism and disposition : the biological fate of chemicals},
  publisher = {American Society for Pharmacology and Experimental Therapeutics},
  address   = {Bethesda},
  issn      = {0090-9556},
  doi       = {10.1124/dmd.115.068395},
  pages     = {1277 -- 1285},
  year      = {2016},
  abstract  = {Aldehyde oxidase (AOX1) is an enzyme with broad substrate specificity, catalyzing the oxidation of a wide range of endogenous and exogenous aldehydes as well as N-heterocyclic aromatic compounds. In humans, the enzyme's role in phase I drug metabolism has been established and its importance is now emerging. However, the true physiologic function of AOX1 in mammals is still unknown. Further, numerous single-nucleotide polymorphisms (SNPs) have been identified in human AOX1. SNPs are a major source of interindividual variability in the human population, and SNP-based amino acid exchanges in AOX1 reportedly modulate the catalytic function of the enzyme in either a positive or negative fashion. For the reliable analysis of the effect of amino acid exchanges in human proteins, the existence of reproducible expression systems for the production of active protein in ample amounts for kinetic, spectroscopic, and crystallographic studies is required. In our study we report an optimized expression system for hAOX1 in Escherichia coli using a codon-optimized construct. The codon-optimization resulted in an up to 15-fold increase of protein production and a simplified purification procedure. The optimized expression system was used to study three SNPs that result in amino acid changes C44W, G1269R, and S1271L. In addition, the crystal structure of the S1271L SNP was solved. We demonstrate that the recombinant enzyme can be used for future studies to exploit the role of AOX in drug metabolism, and for the identification and synthesis of new drugs targeting AOX when combined with crystallographic and modeling studies.},
  language  = {en}
}
@article{SarauliBorowskiPetersetal.2016,
  author    = {Sarauli, David and Borowski, Anja and Peters, Kristina and Schulz, Burkhard and Fattakhova-Rohlfing, Dina and Leimk{\"u}hler, Silke and Lisdat, Fred},
  title     = {Investigation of the pH-Dependent Impact of Sulfonated Polyaniline on Bioelectrocatalytic Activity of Xanthine Dehydrogenase},
  series = {ACS catalysis},
  volume    = {6},
  journal   = {ACS catalysis},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {2155-5435},
  doi       = {10.1021/acscatal.6b02011},
  pages     = {7152 -- 7159},
  year      = {2016},
  abstract  = {We report on the pH-dependent bioelectrocatalytic activity of the redox enzyme xanthine dehydrogenase (XDH) in the presence of sulfonated polyaniline PMSA1 (poly(2-methoxyaniline-5-sulfonic acid)-co-aniline). Ultraviolet-visible (UV-vis) spectroscopic measurements with both components in solution reveal electron transfer from the hypoxanthine (HX)-reduced enzyme to the polymer. The enzyme shows bioelectrocatalytic activity on indium tin oxide (ITO) electrodes, when the polymer is present. Depending on solution pH, different processes can be identified. It can be demonstrated that not only product-based communication with the electrode but also efficient polymer-supported bioelectrocatalysis occur. Interestingly, substrate dependent catalytic currents can be obtained in acidic and neutral solutions, although the highest activity of XDH with natural reaction partners is in the alkaline region. Furthermore, operation of the enzyme electrode without addition of the natural cofactor of XDH is feasible. Finally, macroporous ITO electrodes have been used as an immobilization platform for the fabrication of HX-sensitive electrodes. The study shows that the efficient polymer/enzyme interaction can be advantageously combined with the open structure of an electrode material of controlled pore size, resulting in good processability, stability, and defined signal transfer in the presence of a substrate.},
  language  = {en}
}
@article{ZengFrascaRumschoetteletal.2016,
  author    = {Zeng, Ting and Frasca, Stefano and Rumsch{\"o}ttel, Jens and Koetz, Joachim and Leimk{\"u}hler, Silke and Wollenberger, Ursula},
  title     = {Role of Conductive Nanoparticles in the Direct Unmediated Bioelectrocatalysis of Immobilized Sulfite Oxidase},
  series = {Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis},
  volume    = {28},
  journal   = {Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1040-0397},
  doi       = {10.1002/elan.201600246},
  pages     = {2303 -- 2310},
  year      = {2016},
  language  = {en}
}
@article{CorreiaOtreloCardosoSchwuchowetal.2016,
  author    = {Correia, Marcia A. S. and Otrelo-Cardoso, Ana Rita and Schwuchow, Viola and Clauss, Kajsa G. V. Sigfridsson and Haumann, Michael and Romao, Maria Joao and Leimk{\"u}hler, Silke and Santos-Silva, Teresa},
  title     = {The Escherichia coli Periplasmic Aldehyde Oxidoreductase Is an Exceptional Member of the Xanthine Oxidase Family of Molybdoenzymes},
  series = {ACS chemical biology},
  volume    = {11},
  journal   = {ACS chemical biology},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {1554-8929},
  doi       = {10.1021/acschembio.6b00572},
  pages     = {2923 -- 2935},
  year      = {2016},
  abstract  = {The xanthine oxidase (XO) family comprises molybdenum-dependent enzymes that usually form homodimers (or dimers of heterodimers/trimers) organized in three domains that harbor two [2Fe-2S] clusters, one FAD, and a Mo cofactor. In this work, we crystallized an unusual member of the family, the periplasmic aldehyde oxidoreductase PaoABC from Escherichia coli. This is the first example of an E. coli protein containing a molybdopterin-cytosine-dinucleotide cofactor and is the only heterotrimer of the XO family so far structurally characterized. The crystal structure revealed the presence of an unexpected [4Fe-4S] cluster, anchored to an additional 40 residues subdomain. According to phylogenetic analysis, proteins containing this cluster are widely spread in many bacteria phyla, putatively through repeated gene transfer events. The active site of PaoABC is highly exposed to the surface with no aromatic residues and an arginine (PaoC-R440) making a direct interaction with PaoC-E692, which acts as a base catalyst. In order to understand the importance of R440, kinetic assays were carried out, and the crystal structure of the PaoC-R440H variant was also determined.},
  language  = {en}
}
@article{HanLiOeneretal.2016,
  author    = {Han, Xiao Xia and Li, Junbo and {\"O}ner, Ibrahim Halil and Zhao, Bing and Leimk{\"u}hler, Silke and Hildebrandt, Peter and Weidinger, Inez M.},
  title     = {Nickel electrodes as a cheap and versatile platform for studying structure and function of immobilized redox proteins},
  series = {Analytica chimica acta : an international journal devoted to all branches of analytical chemistry},
  volume    = {941},
  journal   = {Analytica chimica acta : an international journal devoted to all branches of analytical chemistry},
  publisher = {Elsevier},
  address   = {Amsterdam},
  issn      = {0003-2670},
  doi       = {10.1016/j.aca.2016.08.053},
  pages     = {35 -- 40},
  year      = {2016},
  abstract  = {Practical use of many bioelectronic and bioanalytical devices is limited by the need of expensive materials and time consuming fabrication. Here we demonstrate the use of nickel electrodes as a simple and cheap solid support material for bioelectronic applications. The naturally nanostructured electrodes showed a surprisingly high electromagnetic surface enhancement upon light illumination such that immobilization and electron transfer reactions of the model redox proteins cytochrome b(5) (Cyt b(5)) and cytochrome c (Cyt c) could be followed via surface enhanced resonance Raman spectroscopy. It could be shown that the nickel surface, when used as received, promotes a very efficient binding of the proteins upon preservation of their native structure. The immobilized redox proteins could efficiently exchange electrons with the electrode and could even act as an electron relay between the electrode and solubilized myoglobin. Our results open up new possibility for nickel electrodes as an exceptional good support for bioelectronic devices and biosensors on the one hand and for surface enhanced spectroscopic investigations on the other hand. (C) 2016 Elsevier B.V. All rights reserved.},
  language  = {en}
}
@article{CazellesLalaouiHartmannetal.2016,
  author    = {Cazelles, R. and Lalaoui, N. and Hartmann, Tobias and Leimk{\"u}hler, Silke and Wollenberger, Ursula and Antonietti, Markus and Cosnier, S.},
  title     = {Ready to use bioinformatics analysis as a tool to predict immobilisation strategies for protein direct electron transfer (DET)},
  series = {Polymer : the international journal for the science and technology of polymers},
  volume    = {85},
  journal   = {Polymer : the international journal for the science and technology of polymers},
  publisher = {Elsevier},
  address   = {Oxford},
  issn      = {0956-5663},
  doi       = {10.1016/j.bios.2016.04.078},
  pages     = {90 -- 95},
  year      = {2016},
  language  = {en}
}
@misc{RiedelSiemiatkowskaWatanabeetal.2019,
  author    = {Riedel, Simona and Siemiatkowska, Beata and Watanabe, Mutsumi and M{\"u}ller, Christina S. and Sch{\"u}nemann, Volker and Hoefgen, Rainer and Leimk{\"u}hler, Silke},
  title     = {The ABCB7-Like Transporter PexA in Rhodobacter capsulatus Is Involved in the Translocation of Reactive Sulfur Species},
  series = {Postprints der Universit{\"a}t Potsdam Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Postprints der Universit{\"a}t Potsdam Mathematisch-Naturwissenschaftliche Reihe},
  number    = {740},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-43497},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-434975},
  pages     = {10},
  year      = {2019},
  abstract  = {The mitochondrial ATP-binding cassette (ABC) transporters ABCB7 in humans, Atm1 in yeast and ATM3 in plants, are highly conserved in their overall architecture and particularly in their glutathione binding pocket located within the transmembrane spanning domains. These transporters have attracted interest in the last two decades based on their proposed role in connecting the mitochondrial iron sulfur (Fe-S) cluster assembly with its cytosolic Fe-S cluster assembly (CIA) counterpart. So far, the specific compound that is transported across the membrane remains unknown. In this report we characterized the ABCB7-like transporter Rcc02305 in Rhodobacter capsulatus, which shares 47\% amino acid sequence identity with its mitochondrial counterpart. The constructed interposon mutant strain in R. capsulatus displayed increased levels of intracellular reactive oxygen species without a simultaneous accumulation of the cellular iron levels. The inhibition of endogenous glutathione biosynthesis resulted in an increase of total glutathione levels in the mutant strain. Bioinformatic analysis of the amino acid sequence motifs revealed a potential aminotransferase class-V pyridoxal-50-phosphate (PLP) binding site that overlaps with the Walker A motif within the nucleotide binding domains of the transporter. PLP is a well characterized cofactor of L-cysteine desulfurases like IscS and NFS1 which has a role in the formation of a protein-bound persulfide group within these proteins. We therefore suggest renaming the ABCB7-like transporter Rcc02305 in R. capsulatus to PexA for PLP binding exporter. We further suggest that this ABC-transporter in R. capsulatus is involved in the formation and export of polysulfide species to the periplasm.},
  language  = {en}
}
@article{RiedelSiemiatkowskaWatanabeetal.2019,
  author    = {Riedel, Simona and Siemiatkowska, Beata and Watanabe, Mutsumi and M{\"u}ller, Christina S. and Sch{\"u}nemann, Volker and Hoefgen, Rainer and Leimk{\"u}hler, Silke},
  title     = {The ABCB7-Like Transporter PexA in Rhodobacter capsulatus Is Involved in the Translocation of Reactive Sulfur Species},
  series = {Frontiers in Microbiology},
  volume    = {10},
  journal   = {Frontiers in Microbiology},
  publisher = {Frontiers Media},
  address   = {Lausanne},
  issn      = {1664-302X},
  doi       = {10.3389/fmicb.2019.00406},
  pages     = {19},
  year      = {2019},
  abstract  = {The mitochondrial ATP-binding cassette (ABC) transporters ABCB7 in humans, Atm1 in yeast and ATM3 in plants, are highly conserved in their overall architecture and particularly in their glutathione binding pocket located within the transmembrane spanning domains. These transporters have attracted interest in the last two decades based on their proposed role in connecting the mitochondrial iron sulfur (Fe-S) cluster assembly with its cytosolic Fe-S cluster assembly (CIA) counterpart. So far, the specific compound that is transported across the membrane remains unknown. In this report we characterized the ABCB7-like transporter Rcc02305 in Rhodobacter capsulatus, which shares 47\% amino acid sequence identity with its mitochondrial counterpart. The constructed interposon mutant strain in R. capsulatus displayed increased levels of intracellular reactive oxygen species without a simultaneous accumulation of the cellular iron levels. The inhibition of endogenous glutathione biosynthesis resulted in an increase of total glutathione levels in the mutant strain. Bioinformatic analysis of the amino acid sequence motifs revealed a potential aminotransferase class-V pyridoxal-50-phosphate (PLP) binding site that overlaps with the Walker A motif within the nucleotide binding domains of the transporter. PLP is a well characterized cofactor of L-cysteine desulfurases like IscS and NFS1 which has a role in the formation of a protein-bound persulfide group within these proteins. We therefore suggest renaming the ABCB7-like transporter Rcc02305 in R. capsulatus to PexA for PLP binding exporter. We further suggest that this ABC-transporter in R. capsulatus is involved in the formation and export of polysulfide species to the periplasm.},
  language  = {en}
}
@article{McKennaLeimkuehlerHerteretal.2015,
  author    = {McKenna, Shane M. and Leimk{\"u}hler, Silke and Herter, Susanne and Turner, Nicholas J. and Carnell, Andrew J.},
  title     = {Enzyme cascade reactions: synthesis of furandicarboxylic acid (FDCA) and carboxylic acids using oxidases in tandem},
  series = {Green chemistry : an international journal and green chemistry resource},
  volume    = {17},
  journal   = {Green chemistry : an international journal and green chemistry resource},
  number    = {6},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  issn      = {1463-9262},
  doi       = {10.1039/c5gc00707k},
  pages     = {3271 -- 3275},
  year      = {2015},
  abstract  = {A one-pot tandem enzyme reaction using galactose oxidase M3-5 and aldehyde oxidase PaoABC was used to convert hydroxymethylfurfural (HMF) to the pure bioplastics precursor FDCA in 74\% isolated yield. A range of alcohols was also converted to carboxylic acids in high yield under mild conditions.},
  language  = {en}
}
@misc{MendelLeimkuehler2015,
  author    = {Mendel, Ralf R. and Leimk{\"u}hler, Silke},
  title     = {The biosynthesis of the molybdenum cofactors},
  series = {Journal of biological inorganic chemistry},
  volume    = {20},
  journal   = {Journal of biological inorganic chemistry},
  number    = {2},
  publisher = {Springer},
  address   = {New York},
  issn      = {0949-8257},
  doi       = {10.1007/s00775-014-1173-y},
  pages     = {337 -- 347},
  year      = {2015},
  abstract  = {The biosynthesis of the molybdenum cofactors (Moco) is an ancient, ubiquitous, and highly conserved pathway leading to the biochemical activation of molybdenum. Moco is the essential component of a group of redox enzymes, which are diverse in terms of their phylogenetic distribution and their architectures, both at the overall level and in their catalytic geometry. A wide variety of transformations are catalyzed by these enzymes at carbon, sulfur and nitrogen atoms, which include the transfer of an oxo group or two electrons to or from the substrate. More than 50 molybdoenzymes were identified to date. In all molybdoenzymes except nitrogenase, molybdenum is coordinated to a dithiolene group on the 6-alkyl side chain of a pterin called molybdopterin (MPT). The biosynthesis of Moco can be divided into three general steps, with a fourth one present only in bacteria and archaea: (1) formation of the cyclic pyranopterin monophosphate, (2) formation of MPT, (3) insertion of molybdenum into molybdopterin to form Moco, and (4) additional modification of Moco in bacteria with the attachment of a nucleotide to the phosphate group of MPT, forming the dinucleotide variant of Moco. This review will focus on the biosynthesis of Moco in bacteria, humans and plants.},
  language  = {en}
}
@article{ContinFrascaVivekananthanetal.2015,
  author    = {Contin, Andrea and Frasca, Stefano and Vivekananthan, Jeevanthi and Leimk{\"u}hler, Silke and Wollenberger, Ursula and Plumere, Nicolas and Schuhmann, Wolfgang},
  title     = {A pH Responsive Redox Hydrogel for Electrochemical Detection of Redox Silent Biocatalytic Processes. Control of Hydrogel Solvation},
  series = {Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis},
  volume    = {27},
  journal   = {Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis},
  number    = {4},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1040-0397},
  doi       = {10.1002/elan.201400621},
  pages     = {938 -- 944},
  year      = {2015},
  abstract  = {The control of bioelectrocatalytic processes by external stimuli for the indirect detection of non-redox active species was achieved using an esterase and a redox enzyme both integrated within a redox hydrogel. The poly( vinyl) imidazole Os(bpy)(2)Cl hydrogel displays pH-responsive properties. The esterase catalysed reaction leads to a local pH decrease causing protonation of imidazole moieties thus increasing hydrogel solvation and mobility of the tethered Os-complexes. This is the key step to enable improved electron transfer between an aldehyde oxidoreductase and the polymer-bound Os-complexes. The off-on switch is further integrated in a biofuel cell system for self-powered signal generation.},
  language  = {en}
}
@article{SchrapersHartmannKositzkietal.2015,
  author    = {Schrapers, Peer and Hartmann, Tobias and Kositzki, Ramona and Dau, Holger and Reschke, Stefan and Schulzke, Carola and Leimk{\"u}hler, Silke and Haumann, Michael},
  title     = {'Sulfido and Cysteine Ligation Changes at the Molybdenum Cofactor during Substrate Conversion by Formate Dehydrogenase (FDH) from Rhodobacter capsulatus},
  series = {Inorganic chemistry},
  volume    = {54},
  journal   = {Inorganic chemistry},
  number    = {7},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {0020-1669},
  doi       = {10.1021/ic502880y},
  pages     = {3260 -- 3271},
  year      = {2015},
  abstract  = {Formate dehydrogenase (FDH) enzymes are attractive catalysts for potential carbon dioxide conversion applications. The FDH from Rhodobacter capsulatus (RcFDH) binds a bis-molybdopterin-guanine-dinucleotide (bis-MGD) cofactor, facilitating reversible formate (HCOO-) to CO2 oxidation. We characterized the molecular structure of the active site of wildtype RcFDH and protein variants using X-ray absorption spectroscopy (XAS) at the Mo K-edge. This approach has revealed concomitant binding of a sulfido ligand (Mo=S) and a conserved cysteine residue (S(Cys386)) to Mo(VI) in the active oxidized molybdenum cofactor (Moco), retention of such a coordination motif at Mo(V) in a chemically reduced enzyme, and replacement of only the S(Cys386) ligand by an oxygen of formate upon Mo(IV) formation. The lack of a Mo=S bond in RcFDH expressed in the absence of FdsC implies specific metal sulfuration by this bis-MGD binding chaperone. This process still functioned in the Cys386Ser variant, showing no Mo-S(Cys386) ligand, but retaining a Mo=S bond. The C386S variant and the protein expressed without FdsC were inactive in formate oxidation, supporting that both Moligands are essential for catalysis. Low-pH inhibition of RcFDH was attributed to protonation at the conserved His387, supported by the enhanced activity of the His387Met variant at low pH, whereas inactive cofactor species showed sulfido-to-oxo group exchange at the Mo ion. Our results support that the sulfido and S(Cys386) ligands at Mo and a hydrogen-bonded network including His387 are crucial for positioning, deprotonation, and oxidation of formate during the reaction cycle of RcFDH.},
  language  = {en}
}
@article{ZengPankratovFalketal.2015,
  author    = {Zeng, Ting and Pankratov, Dmitry and Falk, Magnus and Leimk{\"u}hler, Silke and Shleev, Sergey and Wollenberger, Ursula},
  title     = {Miniature direct electron transfer based sulphite/oxygen enzymatic fuel cells},
  series = {Biosensors and bioelectronics : the principal international journal devoted to research, design development and application of biosensors and bioelectronics},
  volume    = {66},
  journal   = {Biosensors and bioelectronics : the principal international journal devoted to research, design development and application of biosensors and bioelectronics},
  publisher = {Elsevier},
  address   = {Oxford},
  issn      = {0956-5663},
  doi       = {10.1016/j.bios.2014.10.080},
  pages     = {39 -- 42},
  year      = {2015},
  abstract  = {A direct electron transfer (DET) based sulphite/oxygen biofuel cell is reported that utilises human sulphite oxidase (hSOx) and Myrothecium verrucaria bilirubin oxidase (MvBOx) and nanostructured gold electrodes. For bioanode construction, the nanostructured gold microelectrodes were further modified with 3,3'-dithiodipropionic acid di(N-hydroxysuccinimide ester) to which polyethylene imine was covalently attached. hSOx was adsorbed onto this chemically modified nanostructured electrode with high surface loading of electroactive enzyme and in presence of sulphite high anodic bioelectrocatalytic currents were generated with an onset potential of 0.05 V vs. NHE. The biocathode contained MyBOx directly adsorbed to the deposited gold nanoparticles for cathodic oxygen reduction starting at 0.71 V vs. NHE. Both enzyme electrodes were integrated to a DET-type biofuel cell. Power densities of 8 and 1 mu W cm(-2) were achieved at 0.15 V and 0.45 V of cell voltages, respectively, with the membrane based biodevices under aerobic conditions. (C) 2014 Elsevier B.V. All rights reserved.},
  language  = {en}
}
@misc{YokoyamaLeimkuehler2015,
  author    = {Yokoyama, Kenichi and Leimk{\"u}hler, Silke},
  title     = {The role of FeS clusters for molybdenum cofactor biosynthesis and molybdoenzymes in bacteria},
  series = {Biochimica et biophysica acta : Molecular cell research},
  volume    = {1853},
  journal   = {Biochimica et biophysica acta : Molecular cell research},
  number    = {6},
  publisher = {Elsevier},
  address   = {Amsterdam},
  issn      = {0167-4889},
  doi       = {10.1016/j.bbamcr.2014.09.021},
  pages     = {1335 -- 1349},
  year      = {2015},
  abstract  = {The biosynthesis of the molybdenum cofactor (Moco) has been intensively studied, in addition to its insertion into molybdoenzymes. In particular, a link between the assembly of molybdoenzymes and the biosynthesis of FeS clusters has been identified in the recent years: 1) the synthesis of the first intermediate in Moco biosynthesis requires an FeS-cluster containing protein, 2) the sulfurtransferase for the dithiolene group in Moco is also involved in the synthesis of FeS clusters, thiamin and thiolated tRNAs, 3) the addition of a sulfido-ligand to the molybdenum atom in the active site additionally involves a sulfurtransferase, and 4) most molybdoenzymes in bacteria require FeS clusters as redox active cofactors. In this review we will focus on the biosynthesis of the molybdenum cofactor in bacteria, its modification and insertion into molybdoenzymes, with an emphasis to its link to FeS cluster biosynthesis and sulfur transfer. (C) 2014 Elsevier B.V. All rights reserved.},
  language  = {en}
}
@article{SpricigoLeimkuehlerGortonetal.2015,
  author    = {Spricigo, Roberto and Leimk{\"u}hler, Silke and Gorton, Lo and Scheller, Frieder W. and Wollenberger, Ursula},
  title     = {The Electrically Wired Molybdenum Domain of Human Sulfite Oxidase is Bioelectrocatalytically Active},
  series = {European journal of inorganic chemistry : a journal of ChemPubSoc Europe},
  journal   = {European journal of inorganic chemistry : a journal of ChemPubSoc Europe},
  number    = {21},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1434-1948},
  doi       = {10.1002/ejic.201500034},
  pages     = {3526 -- 3531},
  year      = {2015},
  abstract  = {We report electron transfer between the catalytic molybdenum cofactor (Moco) domain of human sulfite oxidase (hSO) and electrodes through a poly(vinylpyridine)-bound [osmium(N,N'-methyl-2,2'-biimidazole)(3)](2+/3+) complex as the electron-transfer mediator. The biocatalyst was immobilized in this low-potential redox polymer on a carbon electrode. Upon the addition of sulfite to the immobilized separate Moco domain, the generation of a significant catalytic current demonstrated that the catalytic center is effectively wired and active. The bioelectrocatalytic current of the wired separate catalytic domain reached 25\% of the signal of the wired full molybdoheme enzyme hSO, in which the heme b(5) is involved in the electron-transfer pathway. This is the first report on a catalytically active wired molybdenum cofactor domain. The formal potential of this electrochemical mediator is between the potentials of the two cofactors of hSO, and as hSO can occupy several conformations in the polymer matrix, it is imaginable that electron transfer from the catalytic site to the electrode through the osmium center occurs for the hSO molecules in which the Moco domain is sufficiently accessible. The observation of catalytic oxidation currents at low potentials is favorable for applications in bioelectronic devices.},
  language  = {en}
}
@article{HerterMcKennaFrazeretal.2015,
  author    = {Herter, Susanne and McKenna, Shane M. and Frazer, Andrew R. and Leimk{\"u}hler, Silke and Carnell, Andrew J. and Turner, Nicholas J.},
  title     = {Galactose Oxidase Variants for the Oxidation of Amino Alcohols in Enzyme Cascade Synthesis},
  series = {ChemCatChem : heterogeneous \& homogeneous \& bio- \& nano-catalysis ; a journal of ChemPubSoc Europe},
  volume    = {7},
  journal   = {ChemCatChem : heterogeneous \& homogeneous \& bio- \& nano-catalysis ; a journal of ChemPubSoc Europe},
  number    = {15},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1867-3880},
  doi       = {10.1002/cctc.201500218},
  pages     = {2313 -- 2317},
  year      = {2015},
  abstract  = {The use of selected engineered galactose oxidase (GOase) variants for the oxidation of amino alcohols to aldehydes under mild conditions in aqueous systems is reported. GOase variant F-2 catalyses the regioselective oxidation of N-carbobenzyloxy (Cbz)-protected 3-amino-1,2-propanediol to the corresponding -hydroxyaldehyde which was then used in an aldolase reaction. Another variant, M3-5, was found to exhibit activity towards free and N-Cbz-protected aliphatic and aromatic amino alcohols allowing the synthesis of lactams such as 3,4-dihydronaphthalen-1(2H)-one, 2-pyrrolidone and valerolactam in one-pot tandem reactions with xanthine dehydrogenase (XDH) or aldehyde oxidase (PaoABC).},
  language  = {en}
}
@misc{HartmannSchwanholdLeimkuehler2015,
  author    = {Hartmann, Tobias and Schwanhold, Nadine and Leimk{\"u}hler, Silke},
  title     = {Assembly and catalysis of molybdenum or tungsten-containing formate dehydrogenases from bacteria},
  series = {Biochimica et biophysica acta : Proteins and proteomics},
  volume    = {1854},
  journal   = {Biochimica et biophysica acta : Proteins and proteomics},
  number    = {9},
  publisher = {Elsevier},
  address   = {Amsterdam},
  issn      = {1570-9639},
  doi       = {10.1016/j.bbapap.2014.12.006},
  pages     = {1090 -- 1100},
  year      = {2015},
  abstract  = {The global carbon cycle depends on the biological transformations of C-1 compounds, which include the reductive incorporation of CO2 into organic molecules (e.g. in photosynthesis and other autotrophic pathways), in addition to the production of CO2 from formate, a reaction that is catalyzed by formate dehydrogenases (FDHs). FDHs catalyze, in general, the oxidation of formate to CO2 and H+. However, selected enzymes were identified to act as CO2 reductases, which are able to reduce CO2 to formate under physiological conditions. This reaction is of interest for the generation of formate as a convenient storage form of H-2 for future applications. Cofactor-containing FDHs are found in anaerobic bacteria and archaea, in addition to facultative anaerobic or aerobic bacteria. These enzymes are highly diverse and employ different cofactors such as the molybdenum cofactor (Moco), FeS clusters and flavins, or cytochromes. Some enzymes include tungsten (W) in place of molybdenum (Mo) at the active site. For catalytic activity, a selenocysteine (SeCys) or cysteine (Cys) ligand at the Mo atom in the active site is essential for the reaction. This review will focus on the characterization of Mo- and W-containing FDHs from bacteria, their active site structure, subunit compositions and its proposed catalytic mechanism. We will give an overview on the different mechanisms of substrate conversion available so far, in addition to providing an outlook on bio-applications of FDHs. This article is part of a Special Issue entitled: Cofactor-dependent proteins: evolution, chemical diversity and bio-applications. (C) 2014 Elsevier B.V. All rights reserved.},
  language  = {en}
}
@article{HahnEngelhardReschkeetal.2015,
  author    = {Hahn, Aaron and Engelhard, Christopher and Reschke, Stefan and Teutloff, Christian and Bittl, Robert and Leimk{\"u}hler, Silke and Risse, Thomas},
  title     = {Structural Insights into the Incorporation of the Mo Cofactor into Sulfite Oxidase from Site-Directed Spin Labeling},
  series = {Angewandte Chemie : a journal of the Gesellschaft Deutscher Chemiker ; International edition},
  volume    = {54},
  journal   = {Angewandte Chemie : a journal of the Gesellschaft Deutscher Chemiker ; International edition},
  number    = {40},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1433-7851},
  doi       = {10.1002/anie.201504772},
  pages     = {11865 -- 11869},
  year      = {2015},
  abstract  = {Mononuclear molybdoenzymes catalyze a broad range of redox reactions and are highly conserved in all kingdoms of life. This study addresses the question of how the Mo cofactor (Moco) is incorporated into the apo form of human sulfite oxidase (hSO) by using site-directed spin labeling to determine intramolecular distances in the nanometer range. Comparative measurements of the holo and apo forms of hSO enabled the localization of the corresponding structural changes, which are localized to a short loop (residues 263-273) of the Moco-containing domain. A flap-like movement of the loop provides access to the Moco binding-pocket in the apo form of the protein and explains the earlier studies on the in vitro reconstitution of apo-hSO with Moco. Remarkably, the loop motif can be found in a variety of structurally similar molybdoenzymes among various organisms, thus suggesting a common mechanism of Moco incorporation.},
  language  = {en}
}
@article{ZengLeimkuehlerKoetzetal.2015,
  author    = {Zeng, Ting and Leimk{\"u}hler, Silke and Koetz, Joachim and Wollenberger, Ursula},
  title     = {Effective Electrochemistry of Human Sulfite Oxidase Immobilized on Quantum-Dots-Modified Indium Tin Oxide Electrode},
  series = {ACS applied materials \& interfaces},
  volume    = {7},
  journal   = {ACS applied materials \& interfaces},
  number    = {38},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {1944-8244},
  doi       = {10.1021/acsami.5b06665},
  pages     = {21487 -- 21494},
  year      = {2015},
  abstract  = {The bioelectrocatalytic sulfite oxidation by human sulfite oxidase (hSO) on indium tin oxide (ITO) is reported, which is facilitated by functionalizing of the electrode surface with polyethylenimine (PEI)-entrapped CdS nanoparticles and enzyme. hSO was assembled onto the electrode with a high surface loading of electroactive enzyme. In the presence of sulfite but without additional mediators, a high bioelectrocatalytic current was generated. Reference experiments with only PEI showed direct electron transfer and catalytic activity of hSO, but these were less pronounced. The application of the polyelectrolyte-entrapped quantum dots (QDs) on ITO electrodes provides a compatible surface for enzyme binding with promotion of electron transfer. Variations of the buffer solution conditions, e.g., ionic strength, pH, viscosity, and the effect of oxygen, were studied in order to understand intramolecular and heterogeneous electron transfer from hSO to the electrode. The results are consistent with a model derived for the enzyme by using flash photolysis in solution and spectroelectrochemistry and molecular dynamic simulations of hSO on monolayer-modified gold electrodes. Moreover, for the first time a photoelectrochemical electrode involving immobilized hSO is demonstrated where photoexcitation of the CdS/hSO-modified electrode lead to an enhanced generation of bioelectrocatalytic currents upon sulfite addition. Oxidation starts already at the redox potential of the electron transfer domain of hSO and is greatly increased by application of a small overpotential to the CdS/hSO-modified ITO.},
  language  = {en}
}
@article{CoelhoFotiHartmannetal.2015,
  author    = {Coelho, Catarina and Foti, Alessandro and Hartmann, Tobias and Santos-Silva, Teresa and Leimk{\"u}hler, Silke and Romao, Maria Joao},
  title     = {Structural insights into xenobiotic and inhibitor binding to human aldehyde oxidase},
  series = {Nature chemical biology},
  volume    = {11},
  journal   = {Nature chemical biology},
  number    = {10},
  publisher = {Nature Publ. Group},
  address   = {New York},
  issn      = {1552-4450},
  doi       = {10.1038/NCHEMBIO.1895},
  pages     = {779 -- +},
  year      = {2015},
  abstract  = {Aldehyde oxidase (AOX) is a xanthine oxidase (XO)-related enzyme with emerging importance due to its role in the metabolism of drugs and xenobiotics. We report the first crystal structures of human AOX1, substrate free (2.6-angstrom resolution) and in complex with the substrate phthalazine and the inhibitor thioridazine (2.7-angstrom resolution). Analysis of the protein active site combined with steady-state kinetic studies highlight the unique features, including binding and substrate orientation at the active site, that characterize human AOX1 as an important drug-metabolizing enzyme. Structural analysis of the complex with the noncompetitive inhibitor thioridazine revealed a new, unexpected and fully occupied inhibitor-binding site that is structurally conserved among mammalian AOXs and XO. The new structural insights into the catalytic and inhibition mechanisms of human AOX that we now report will be of great value for the rational analysis of clinical drug interactions involving inhibition of AOX1 and for the prediction and design of AOX-stable putative drugs.},
  language  = {en}
}
@article{OtreloCardosoSchwuchowRodriguesetal.2014,
  author    = {Otrelo-Cardoso, Ana Rita and Schwuchow, Viola and Rodrigues, David and Cabrita, Eurico J. and Leimk{\"u}hler, Silke and Romao, Maria Joao and Santos-Silva, Teresa},
  title     = {Biochemical, stabilization and crystallization studies on a molecular chaperone (PaoD) involved in the maturation of molybdoenzymes},
  series = {PLoS one},
  volume    = {9},
  journal   = {PLoS one},
  number    = {1},
  publisher = {PLoS},
  address   = {San Fransisco},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0087295},
  pages     = {9},
  year      = {2014},
  abstract  = {Molybdenum and tungsten enzymes require specific chaperones for folding and cofactor insertion. PaoD is the chaperone of the periplasmic aldehyde oxidoreductase PaoABC. It is the last gene in the paoABCD operon in Escherichia coli and its presence is crucial for obtaining mature enzyme. PaoD is an unstable, 35 kDa, protein. Our biochemical studies showed that it is a dimer in solution with a tendency to form large aggregates, especially after freezing/thawing cycles. In order to improve stability, PaoD was thawed in the presence of two ionic liquids [C(4)mim]Cl and [C(2)OHmim]PF6 and no protein precipitation was observed. This allowed protein concentration and crystallization using polyethylene glycol or ammonium sulfate as precipitating agents. Saturation transfer difference - nuclear magnetic resonance (STD-NMR) experiments have also been performed in order to investigate the effect of the ionic liquids in the stabilization process, showing a clear interaction between the acidic ring protons of the cation and, most likely, negatively charged residues at the protein surface. DLS assays also show a reduction of the overall size of the protein aggregates in presence of ionic liquids. Furthermore, cofactor binding studies on PaoD showed that the protein is able to discriminate between molybdenum and tungsten bound to the molybdenum cofactor, since only a Mo-MPT form of the cofactor remained bound to PaoD.},
  language  = {en}
}
@article{OtreloCardosodaSilvaCorreiaSchwuchowetal.2014,
  author    = {Otrelo-Cardoso, Ana Rita and da Silva Correia, Marcia Alexandra and Schwuchow, Viola and Svergun, Dmitri I. and Romao, Maria Joao and Leimk{\"u}hler, Silke and Santos-Silva, Teresa},
  title     = {Structural Data on the Periplasmic Aldehyde Oxidoreductase PaoABC from Escherichia coli: SAXS and Preliminary X-ray Crystallography Analysis},
  series = {International journal of molecular sciences},
  volume    = {15},
  journal   = {International journal of molecular sciences},
  number    = {2},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {1422-0067},
  doi       = {10.3390/ijms15022223},
  pages     = {2223 -- 2236},
  year      = {2014},
  abstract  = {The periplasmic aldehyde oxidoreductase PaoABC from Escherichia coli is a molybdenum enzyme involved in detoxification of aldehydes in the cell. It is an example of an heterotrimeric enzyme of the xanthine oxidase family of enzymes which does not dimerize via its molybdenum cofactor binding domain. In order to structurally characterize PaoABC, X-ray crystallography and small angle X-ray scattering (SAXS) have been carried out. The protein crystallizes in the presence of 20\% (w/v) polyethylene glycol 3350 using the hanging-drop vapour diffusion method. Although crystals were initially twinned, several experiments were done to overcome twinning and lowering the crystallization temperature (293 K to 277 K) was the solution to the problem. The non-twinned crystals used to solve the structure diffract X-rays to beyond 1.80 angstrom and belong to the C2 space group, with cell parameters a = 109.42 angstrom, b = 78.08 angstrom, c = 151.77 angstrom, = 99.77 degrees, and one molecule in the asymmetric unit. A molecular replacement solution was found for each subunit separately, using several proteins as search models. SAXS data of PaoABC were also collected showing that, in solution, the protein is also an heterotrimer.},
  language  = {en}
}
@article{BoehmerHartmannLeimkuehler2014,
  author    = {Boehmer, Nadine and Hartmann, Tobias and Leimk{\"u}hler, Silke},
  title     = {The chaperone FdsC for Rhodobacter capsulatus formate dehydrogenase binds the bis-molybdopterin guanine dinucleotide cofactor},
  series = {FEBS letters : the journal for rapid publication of short reports in molecular biosciences},
  volume    = {588},
  journal   = {FEBS letters : the journal for rapid publication of short reports in molecular biosciences},
  number    = {4},
  publisher = {Elsevier},
  address   = {Amsterdam},
  issn      = {0014-5793},
  doi       = {10.1016/j.febslet.2013.12.033},
  pages     = {531 -- 537},
  year      = {2014},
  abstract  = {Molybdoenzymes are complex enzymes in which the molybdenum cofactor (Moco) is deeply buried in the enzyme. Most molybdoenzymes contain a specific chaperone for the insertion of Moco. For the formate dehydrogenase FdsGBA from Rhodobacter capsulatus the two chaperones FdsC and FdsD were identified to be essential for enzyme activity, but are not a subunit of the mature enzyme. Here, we purified and characterized the FdsC protein after heterologous expression in Escherichia coli. We were able to copurify FdsC with the bound Moco derivate bis-molybdopterin guanine dinucleotide. This cofactor successfully was used as a source to reconstitute the activity of molybdoenzymes. Structured summary of protein interactions: FdsC and FdsC bind by molecular sieving (View interaction) FdsD binds to RcMobA by surface plasmon resonance (View interaction) FdsC binds to RcMobA by surface plasmon resonance (View interaction) FdsC binds to FdsA by surface plasmon resonance (View interaction)},
  language  = {en}
}
@inproceedings{Leimkuehler2014,
  author    = {Leimk{\"u}hler, Silke},
  title     = {Studies on the Oxygen tolerant formate deyhdrogenase from rhodobacter capsulatus},
  series = {Journal of biological inorganic chemistry},
  volume    = {19},
  booktitle = {Journal of biological inorganic chemistry},
  publisher = {Springer},
  address   = {New York},
  issn      = {0949-8257},
  pages     = {S72 -- S72},
  year      = {2014},
  language  = {en}
}
@article{MareljaDambowskyBolisetal.2014,
  author    = {Marelja, Zvonimir and Dambowsky, Miriam and Bolis, Marco and Georgiou, Marina L. and Garattini, Enrico and Missirlis, Fanis and Leimk{\"u}hler, Silke},
  title     = {The four aldehyde oxidases of Drosophila melanogaster have different gene expression patterns and enzyme substrate specificities},
  series = {The journal of experimental biology},
  volume    = {217},
  journal   = {The journal of experimental biology},
  number    = {12},
  publisher = {Company of Biologists Limited},
  address   = {Cambridge},
  issn      = {0022-0949},
  doi       = {10.1242/jeb.102129},
  pages     = {2201 -- 2211},
  year      = {2014},
  abstract  = {In the genome of Drosophila melanogaster, four genes coding for aldehyde oxidases (AOX1-4) were identified on chromosome 3. Phylogenetic analysis showed that the AOX gene cluster evolved via independent duplication events in the vertebrate and invertebrate lineages. The functional role and the substrate specificity of the distinct Drosophila AOX enzymes is unknown. Two loss-of-function mutant alleles in this gene region, low pyridoxal oxidase (Po-lpo) and aldehyde oxidase-1 (Aldox-1(n1)) are associated with a phenotype characterized by undetectable AOX enzymatic activity. However, the genes involved and the corresponding mutations have not yet been identified. In this study we characterized the activities, substrate specificities and expression profiles of the four AOX enzymes in D. melanogaster. We show that the Po-lpo-associated phenotype is the consequence of a structural alteration of the AOX1 gene. We identified an 11-bp deletion in the Po-lpo allele, resulting in a frame-shift event, which removes the molybdenum cofactor domain of the encoded enzyme. Furthermore, we show that AOX2 activity is detectable only during metamorphosis and characterize a Minos-AOX2 insertion in this developmental gene that disrupts its activity. We demonstrate that the Aldox-1(n1) phenotype maps to the AOX3 gene and AOX4 activity is not detectable in our assays.},
  language  = {en}
}
@article{DeyAdamovskiFriebeetal.2014,
  author    = {Dey, Pradip and Adamovski, Miriam and Friebe, Simon and Badalyan, Artavazd and Mutihac, Radu-Cristian and Paulus, Florian and Leimk{\"u}hler, Silke and Wollenberger, Ursula and Haag, Rainer},
  title     = {Dendritic polyglycerol-poly(ethylene glycol)-based polymer networks for biosensing application},
  series = {ACS applied materials \& interfaces},
  volume    = {6},
  journal   = {ACS applied materials \& interfaces},
  number    = {12},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {1944-8244},
  doi       = {10.1021/am502018x},
  pages     = {8937 -- 8941},
  year      = {2014},
  abstract  = {This work describes the formation of a new dendritic polyglycerol-poly(ethylene glycol)-based 3D polymer network as a matrix for immobilization of the redox enzyme periplasmatic aldehyde oxidoreductase to create an electrochemical biosensor. The novel network is built directly on the gold surface, where it simultaneously stabilizes the enzyme for up to 4 days. The prepared biosensors can be used for amperometric detection of benzaldehyde in the range of 0.8-400 mu M.},
  language  = {en}
}
@article{HahnReschkeLeimkuehleretal.2014,
  author    = {Hahn, Aaron and Reschke, Stefan and Leimk{\"u}hler, Silke and Risse, Thomas},
  title     = {Ketoxime coupling of p-Acetylphenylalanine at neutral pH for site-directed spin labeling of human sulfite oxidase},
  series = {The journal of physical chemistry : B, Condensed matter, materials, surfaces, interfaces \& biophysical chemistry},
  volume    = {118},
  journal   = {The journal of physical chemistry : B, Condensed matter, materials, surfaces, interfaces \& biophysical chemistry},
  number    = {25},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {1520-6106},
  doi       = {10.1021/jp503471j},
  pages     = {7077 -- 7084},
  year      = {2014},
  abstract  = {Site-directed spin labeling of the unnatural amino acid p-acetylphenylalanine (p-AcPhe) using oxime based coupling chemistry is successfully applied to investigate human sulfite oxidase (hSO), a protein containing an essential cysteine residue, which impedes the use of thiol based coupling chemistry. The protein was found to be sensitive toward typical reaction conditions of oxime coupling, namely, acidic reaction conditions and elevated temperatures. Thus, coupling at neutral pH and room temperature is mandatory. Three catalysts described in the literature to accelerate the reaction rate have been tested. Best spin labeling efficiencies were observed for p-methoxyaniline, while the other catalysts described in the literature to have even better performance for oxime coupling at neutral pH were substantially less active or led to precipitation of the protein. A clear correlation of spin labeling efficiency with the local environment of the residue is found, shedding some light on the importance of the sterically demanding reaction complex between p-AcPhe, the aniline catalyst, and the spin label for the reaction rate. The analysis of the line shape has shown that its interpretation in terms of local environment is more challenging as compared to the well-established spin labels based on cysteine chemistry. To this end the results presented here indicate that the larger steric demand of the spin labeled p-AcPhe can induce structural effects instead of reporting on them.},
  language  = {en}
}
@article{BechiHerterMcKennaetal.2014,
  author    = {Bechi, Beatrice and Herter, Susanne and McKenna, Shane and Riley, Christopher and Leimk{\"u}hler, Silke and Turner, Nicholas J. and Carnell, Andrew J.},
  title     = {Catalytic bio-chemo and bio-bio tandem oxidation reactions for amide and carboxylic acid synthesis},
  series = {Green chemistry : an international journal and green chemistry resource},
  volume    = {16},
  journal   = {Green chemistry : an international journal and green chemistry resource},
  number    = {10},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  issn      = {1463-9262},
  doi       = {10.1039/c4gc01321b},
  pages     = {4524 -- 4529},
  year      = {2014},
  abstract  = {A catalytic toolbox for three different water-based one-pot cascades to convert aryl alcohols to amides and acids and cyclic amines to lactams, involving combination of oxidative enzymes (monoamine oxidase, xanthine dehydrogenase, galactose oxidase and laccase) and chemical oxidants (TBHP or Cul(cat)/H2O2) at mild temperatures, is presented. Mutually compatible conditions were found to afford products in good to excellent yields.},
  language  = {en}
}
@article{HallReschkeCaoetal.2014,
  author    = {Hall, James and Reschke, Stefan and Cao, Hongnan and Leimk{\"u}hler, Silke and Hille, Russ},
  title     = {The reductive half-reaction of xanthine dehydrogenase from rhodobacter capsulatus the role of GLU(232) in catalysis},
  series = {The journal of biological chemistry},
  volume    = {289},
  journal   = {The journal of biological chemistry},
  number    = {46},
  publisher = {American Society for Biochemistry and Molecular Biology},
  address   = {Bethesda},
  issn      = {0021-9258},
  doi       = {10.1074/jbc.M114.603456},
  pages     = {32121 -- 32130},
  year      = {2014},
  abstract  = {Background: Kinetic characterization of wild-type xanthine dehydrogenase and variants. Results: Comparison of the pH dependence of both k(red) and k(red)/K-d, as well as k(cat) and k(cat)/K-m. Conclusion: Ionized Glu(232) of wild-type enzyme plays an important role in catalysis by discriminating against the monoanionic form of xanthine. Significance: Examining the contributions of Glu(232) to catalysis is essential for understanding the mechanism of xanthine dehydrogenase. The kinetic properties of an E232Q variant of the xanthine dehydrogenase from Rhodobacter capsulatus have been examined to ascertain whether Glu(232) in wild-type enzyme is protonated or unprotonated in the course of catalysis at neutral pH. We find that k(red), the limiting rate constant for reduction at high [xanthine], is significantly compromised in the variant, a result that is inconsistent with Glu(232) being neutral in the active site of the wild-type enzyme. A comparison of the pH dependence of both k(red) and k(red)/K-d from reductive half-reaction experiments between wild-type and enzyme and the E232Q variant suggests that the ionized Glu(232) of wild-type enzyme plays an important role in catalysis by discriminating against the monoanionic form of substrate, effectively increasing the pK(a) of substrate by two pH units and ensuring that at physiological pH the neutral form of substrate predominates in the Michaelis complex. A kinetic isotope study of the wild-type R. capsulatus enzyme indicates that, as previously determined for the bovine and chicken enzymes, product release is principally rate-limiting in catalysis. The disparity in rate constants for the chemical step of the reaction and product release, however, is not as great in the bacterial enzyme as compared with the vertebrate forms. The results indicate that the bacterial and bovine enzymes catalyze the chemical step of the reaction to the same degree and that the faster turnover observed with the bacterial enzyme is due to a faster rate constant for product release than is seen with the vertebrate enzyme.},
  language  = {en}
}
@article{FraesdorfRadonLeimkuehler2014,
  author    = {Fraesdorf, Benjamin and Radon, Christin and Leimk{\"u}hler, Silke},
  title     = {Characterization and interaction studies of two isoforms of the dual localized 3-mercaptopyruvate sulfurtransferase TUM1 from humans},
  series = {The journal of biological chemistry},
  volume    = {289},
  journal   = {The journal of biological chemistry},
  number    = {50},
  publisher = {American Society for Biochemistry and Molecular Biology},
  address   = {Bethesda},
  issn      = {0021-9258},
  doi       = {10.1074/jbc.M114.605733},
  pages     = {34543 -- 34556},
  year      = {2014},
  abstract  = {Background: Localization and identification of interaction partners of two splice variants of the human 3-mercaptopyruvate sulfurtransferase TUM1. Results: We show that TUM1 interacts with proteins involved in Moco and FeS cluster biosynthesis. Conclusion: Human TUM1 is a dual localized protein in the cytosol and mitochondria with distinct roles in sulfur transfer and interaction partners. Significance: The study contributes to the sulfur transfer pathway for the biosynthesis of sulfur-containing biofactors. The human tRNA thiouridine modification protein (TUM1), also designated as 3-mercaptopyruvate sulfurtransferase (MPST), has been implicated in a wide range of physiological processes in the cell. The roles range from an involvement in thiolation of cytosolic tRNAs to the generation of H2S as signaling molecule both in mitochondria and the cytosol. TUM1 is a member of the sulfurtransferase family and catalyzes the conversion of 3-mercaptopyruvate to pyruvate and protein-bound persulfide. Here, we purified and characterized two novel TUM1 splice variants, designated as TUM1-Iso1 and TUM1-Iso2. The purified proteins showed similar kinetic behavior and comparable pH and temperature dependence. Cellular localization studies, however, showed a different localization pattern between the isoforms. TUM1-Iso1 is exclusively localized in the cytosol, whereas TUM1-Iso2 showed a dual localization both in the cytosol and mitochondria. Interaction studies were performed with the isoforms both in vitro using the purified proteins and in vivo by fluorescence analysis in human cells, using the split-EGFP system. The studies showed that TUM1 interacts with the l-cysteine desulfurase NFS1 and the rhodanese-like protein MOCS3, suggesting a dual function of TUM1 both in sulfur transfer for the biosynthesis of the molybdenum cofactor, and for the thiolation of tRNA. Our studies point to distinct roles of each TUM1 isoform in the sulfur transfer processes in the cell, with different compartmentalization of the two splice variants of TUM1.},
  language  = {en}
}
@article{NeumannSedukIobbiNivoletal.2011,
  author    = {Neumann, Meina and Seduk, Farida and Iobbi-Nivol, Chantal and Leimk{\"u}hler, Silke},
  title     = {Molybdopterin Dinucleotide Biosynthesis in Escherichia coli identification of amino acid residues of molybdopterin dinucleotide transferases that determine specificity for binding of guanine or cytosine nucleotides},
  series = {The journal of biological chemistry},
  volume    = {286},
  journal   = {The journal of biological chemistry},
  number    = {2},
  publisher = {American Society for Biochemistry and Molecular Biology},
  address   = {Bethesda},
  issn      = {0021-9258},
  doi       = {10.1074/jbc.M110.155671},
  pages     = {1400 -- 1408},
  year      = {2011},
  abstract  = {The molybdenum cofactor is modified by the addition of GMP or CMP to the C4' phosphate of molybdopterin forming the molybdopterin guanine dinucleotide or molybdopterin cytosine dinucleotide cofactor, respectively. The two reactions are catalyzed by specific enzymes as follows: the GTP: molybdopterin guanylyltransferase MobA and the CTP: molybdopterin cytidylyltransferase MocA. Both enzymes show 22\% amino acid sequence identity and are specific for their respective nucleotides. Crystal structure analysis of MobA revealed two conserved motifs in the N-terminal domain of the protein involved in binding of the guanine base. Based on these motifs, we performed site-directed mutagenesis studies to exchange the amino acids to the sequence found in the paralogue MocA. Using a fully defined in vitro system, we showed that the exchange of five amino acids was enough to obtain activity with both GTP and CTP in either MocA or MobA. Exchange of the complete N-terminal domain of each protein resulted in the total inversion of nucleotide specificity activity, showing that the N-terminal domain determines nucleotide recognition and binding. Analysis of protein-protein interactions showed that the C-terminal domain of either MocA or MobA determines the specific binding to the respective acceptor protein.},
  language  = {en}
}
@article{KalimuthuLeimkuehlerBernhardt2011,
  author    = {Kalimuthu, Palraj and Leimk{\"u}hler, Silke and Bernhardt, Paul V.},
  title     = {Xanthine dehydrogenase electrocatalysis autocatalysis and novel activity},
  series = {The journal of physical chemistry : B, Condensed matter, materials, surfaces, interfaces \& biophysical chemistry},
  volume    = {115},
  journal   = {The journal of physical chemistry : B, Condensed matter, materials, surfaces, interfaces \& biophysical chemistry},
  number    = {11},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {1520-6106},
  doi       = {10.1021/jp111809f},
  pages     = {2655 -- 2662},
  year      = {2011},
  abstract  = {The enzyme xanthine dehydrogenase (XDH) from the purple photosynthetic bacterium Rhodobacter capsulatus catalyzes the oxidation of hypoxanthine to xanthine and xanthine to uric acid as part of purine metabolism. The native electron acceptor is NAD(+) but herein we show that uric acid in its 2-electron oxidized form is able to act as an artificial electron acceptor from XDH in an electrochemically driven catalytic system. Hypoxanthine oxidation is also observed with the novel production of uric acid in a series of two consecutive 2-electron oxidation reactions via xanthine. XDH exhibits native activity in terms of its pH optimum and inhibition by allopurinol.},
  language  = {en}
}
@misc{LeimkuehlerWuebbensRajagopalan2011,
  author    = {Leimk{\"u}hler, Silke and Wuebbens, Margot M. and Rajagopalan, K. V.},
  title     = {The history of the discovery of the molybdenum cofactor and novel aspects of its biosynthesis in bacteria},
  series = {Coordination chemistry reviews},
  volume    = {255},
  journal   = {Coordination chemistry reviews},
  number    = {9-10},
  publisher = {Elsevier},
  address   = {Lausanne},
  issn      = {0010-8545},
  doi       = {10.1016/j.ccr.2010.12.003},
  pages     = {1129 -- 1144},
  year      = {2011},
  abstract  = {The biosynthesis of the molybdenum cofactor in bacteria is described with a detailed analysis of each individual reaction leading to the formation of stable intermediates during the synthesis of molybdopterin from GTP. As a starting point, the discovery of molybdopterin and the elucidation of its structure through the study of stable degradation products are described. Subsequent to molybdopterin synthesis, the molybdenum atom is added to the molybdopterin dithiolene group to form the molybdenum cofactor. This cofactor is either inserted directly into specific molybdoenzymes or is further modified by the addition of nucleotides to molybdopterin phosphate group or the replacement of ligands at the molybdenum center.},
  language  = {en}
}
@article{MahroCoelhoTrincaoetal.2011,
  author    = {Mahro, Martin and Coelho, Catarina and Trincao, Jose and Rodrigues, David and Terao, Mineko and Garattini, Enrico and Saggu, Miguel and Lendzian, Friedhelm and Hildebrandt, Peter and Romao, Maria Joao and Leimk{\"u}hler, Silke},
  title     = {Characterization and crystallization of mouse aldehyde oxidase 3 - from mouse liver to escherichia coli heterologous protein expression},
  series = {Drug metabolism and disposition : the biological fate of chemicals},
  volume    = {39},
  journal   = {Drug metabolism and disposition : the biological fate of chemicals},
  number    = {10},
  publisher = {American Society for Pharmacology and Experimental Therapeutics},
  address   = {Bethesda},
  issn      = {0090-9556},
  doi       = {10.1124/dmd.111.040873},
  pages     = {1939 -- 1945},
  year      = {2011},
  abstract  = {Aldehyde oxidase (AOX) is characterized by a broad substrate specificity, oxidizing aromatic azaheterocycles, such as N(1)-methylnicotinamide and N-methylphthalazinium, or aldehydes, such as benzaldehyde, retinal, and vanillin. In the past decade, AOX has been recognized increasingly to play an important role in the metabolism of drugs through its complex cofactor content, tissue distribution, and substrate recognition. In humans, only one AOX gene (AOX1) is present, but in mouse and other mammals different AOX homologs were identified. The multiple AOX isoforms are expressed tissue-specifically in different organisms, and it is believed that they recognize distinct substrates and carry out different physiological tasks. AOX is a dimer with a molecular mass of approximately 300 kDa, and each subunit of the homodimeric enzyme contains four different cofactors: the molybdenum cofactor, two distinct [2Fe-2S] clusters, and one FAD. We purified the AOX homolog from mouse liver (mAOX3) and established a system for the heterologous expression of mAOX3 in Escherichia coli. The purified enzymes were compared. Both proteins show the same characteristics and catalytic properties, with the difference that the recombinant protein was expressed and purified in a 30\% active form, whereas the native protein is 100\% active. Spectroscopic characterization showed that FeSII is not assembled completely in mAOX3. In addition, both proteins were crystallized. The best crystals were from native mAOX3 and diffracted beyond 2.9 angstrom. The crystals belong to space group P1, and two dimers are present in the unit cell.},
  language  = {en}
}
@article{SamuelHornDoeringetal.2011,
  author    = {Samuel, Prinson P. and Horn, Sebastian and D{\"o}ring, Alexander and Havelius, Kajsa G. V. and Reschke, Stefan and Leimk{\"u}hler, Silke and Haumann, Michael and Schulzke, Carola},
  title     = {A Crystallographic and Mo K-Edge XAS Study of Molybdenum Oxo Bis-,Mono-, and Non-Dithiolene Complexes - First-Sphere Coordination Geometry and Noninnocence of Ligands},
  series = {European journal of inorganic chemistry : a journal of ChemPubSoc Europe},
  journal   = {European journal of inorganic chemistry : a journal of ChemPubSoc Europe},
  number    = {28},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1434-1948},
  doi       = {10.1002/ejic.201100331},
  pages     = {4387 -- 4399},
  year      = {2011},
  abstract  = {Ten square-based pyramidal molybdenum complexes with different sulfur donor ligands, that is, a variety of dithiolenes and sulfides, were prepared, which mimic coordination motifs of the molybdenum cofactors of molybdenum-dependent oxidoreductases. The model compounds were investigated by Mo K-edge X-ray absorption spectroscopy (XAS) and (with one exception) their molecular structures were analyzed by X-ray diffraction to derive detailed information on bond lengths and geometries of the first coordination shell of molybdenum. Only small variations in Mo=O and Mo-S bond lengths and their respective coordination angles were observed for all complexes including those containing Mo(CO)(2) or Mo(mu-S)(2)Mo motifs. XAS analysis (edge energy) revealed higher relative oxidation levels in the molybdenum ion in compounds with innocent sulfur-based ligands relative to those in dithiolene complexes, which are known to exhibit noninnocence, that is, donation of substantial electron density from ligand to metal. In addition, longer average Mo-S and Mo=O bonds and consequently lower.(Mo=O) stretching frequencies in the IR spectra were observed for complexes with dithiolene-derived ligands. The results emphasize that the noninnocent character of the dithiolene ligand influences the electronic structure of the model compounds, but does not significantly affect their metal coordination geometry, which is largely determined by the Mo(IV) or (V) ion itself. The latter conclusion also holds for the molybdenum site geometries in the oxidized Mo-VI cofactor of DMSO reductase and the reduced Mo-IV cofactor of arsenite oxidase. The innocent behavior of the dithiolene molybdopterin ligands observed in the enzymes is likely to be related to cofactor-protein interactions.},
  language  = {en}
}
@article{DahlUrbanBolteetal.2011,
  author    = {Dahl, Jan-Ulrik and Urban, Alexander and Bolte, Andrea and Sriyabhaya, Promjit and Donahue, Janet L. and Nimtz, Manfred and Larson, Timothy J. and Leimk{\"u}hler, Silke},
  title     = {The identification of a novel protein involved in Molybdenum Cofactor Biosynthesis in Escherichia coli},
  series = {The journal of biological chemistry},
  volume    = {286},
  journal   = {The journal of biological chemistry},
  number    = {41},
  publisher = {American Society for Biochemistry and Molecular Biology},
  address   = {Bethesda},
  issn      = {0021-9258},
  doi       = {10.1074/jbc.M111.282368},
  pages     = {35801 -- 35812},
  year      = {2011},
  abstract  = {Background: In Moco biosynthesis, sulfur is transferred from L-cysteine to MPT synthase, catalyzing the conversion of cPMP to MPT. Results: The rhodanese-like protein YnjE is a novel protein involved in Moco biosynthesis. Conclusion: YnjE enhances the rate of conversion of cPMP to MPT and interacts with MoeB and IscS. S ignificance: To understand the mechanism of sulfur transfer and the role of rhodaneses in the cell.},
  language  = {en}
}
@inproceedings{LeimkuehlerHartmannGarattinietal.2011,
  author    = {Leimk{\"u}hler, Silke and Hartmann, Tobias and Garattini, Enrico and Jones, Jeffrey P.},
  title     = {Structure-function studies on human aldehyde oxidase and the impact of polymorphisms on enzyme activity},
  series = {Drug metabolism reviews : biotransformation and disposition of xenobiotics ; official journal of the International Society for the Study of Xenobiotics},
  volume    = {43},
  booktitle = {Drug metabolism reviews : biotransformation and disposition of xenobiotics ; official journal of the International Society for the Study of Xenobiotics},
  number    = {6},
  publisher = {Taylor \& Francis Group},
  address   = {London},
  issn      = {0360-2532},
  pages     = {13 -- 13},
  year      = {2011},
  language  = {en}
}
@article{VossNimtzLeimkuehler2011,
  author    = {Voss, Martin and Nimtz, Manfred and Leimk{\"u}hler, Silke},
  title     = {Elucidation of the dual role of Mycobacterial MoeZR in Molybdenum Cofactor Biosynthesis and Cysteine Biosynthesis},
  series = {PLoS one},
  volume    = {6},
  journal   = {PLoS one},
  number    = {11},
  publisher = {PLoS},
  address   = {San Fransisco},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0028170},
  pages     = {9},
  year      = {2011},
  abstract  = {The pathway of molybdenum cofactor biosynthesis has been studied in detail by using proteins from Mycobacterium species, which contain several homologs associated with the first steps of Moco biosynthesis. While all Mycobacteria species contain a MoeZR, only some strains have acquired an additional homolog, MoeBR, by horizontal gene transfer. The role of MoeBR and MoeZR was studied in detail for the interaction with the two MoaD-homologs involved in Moco biosynthesis, MoaD1 and MoaD2, in addition to the CysO protein involved in cysteine biosynthesis. We show that both proteins have a role in Moco biosynthesis, while only MoeZR, but not MoeBR, has an additional role in cysteine biosynthesis. MoeZR and MoeBR were able to complement an E. coli moeB mutant strain, but only in conjunction with the Mycobacterial MoaD1 or MoaD2 proteins. Both proteins were able to sulfurate MoaD1 and MoaD2 in vivo, while only MoeZR additionally transferred the sulfur to CysO. Our in vivo studies show that Mycobacteria have acquired several homologs to maintain Moco biosynthesis. MoeZR has a dual role in Moco- and cysteine biosynthesis and is involved in the sulfuration of MoaD and CysO, whereas MoeBR only has a role in Moco biosynthesis, which is not an essential function for Mycobacteria.},
  language  = {en}
}