@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} } @article{RedelbergerSedukGenestetal.2011, author = {Redelberger, David and Seduk, Farida and Genest, Olivier and Mejean, Vincent and Leimk{\"u}hler, Silke and Iobbi-Nivol, Chantal}, title = {YcdY Protein of Escherichia coli, an Atypical Member of the TorD Chaperone Family}, series = {Journal of bacteriology}, volume = {193}, journal = {Journal of bacteriology}, number = {23}, publisher = {American Society for Microbiology}, address = {Washington}, issn = {0021-9193}, doi = {10.1128/JB.05927-11}, pages = {6512 -- 6516}, year = {2011}, abstract = {The TorD family of specific chaperones is divided into four subfamilies dedicated to molybdoenzyme biogenesis and a fifth one, exemplified by YcdY of Escherichia coli, for which no defined partner has been identified so far. We propose that YcdY is the chaperone of YcdX, a zinc protein involved in the swarming motility process of E. coli, since YcdY interacts with YcdX and increases its activity in vitro.}, language = {en} }