TY  - GEN
A1  - Tiedemann, Kim
A1  - Iobbi-Nivol, Chantal
A1  - Leimkühler, Silke
T1  - The Role of the Nucleotides in the Insertion of the bis-Molybdopterin Guanine Dinucleotide Cofactor into apo-Molybdoenzymes
T2  - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
N2  - 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.
T3  - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 1268 
KW  - bis-MGD
KW  - chaperone
KW  - molybdenum cofactor
KW  - TMAO reductase
Y1  - 2022
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-561728
SN  - 1866-8372
SP  - 1
EP  - 15
PB  - Universitätsverlag Potsdam
CY  - Potsdam
ER  - 
TY  - JOUR
A1  - Mitrova, Biljana
A1  - Tadjoung Waffo, Armel Franklin
A1  - Kaufmann, Paul
A1  - Iobbi-Nivol, Chantal
A1  - Leimkühler, Silke
A1  - Wollenberger, Ulla
T1  - Trimethylamine N-Oxide Electrochemical Biosensor with a Chimeric Enzyme
JF  - ChemElectroChem
N2  - 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.
KW  - trimethylamine N-oxide (TMAO)
KW  - TMAO reductase
KW  - chimeric enzyme
KW  - molybdoenzyme
KW  - electrochemical biosensor
Y1  - 2018
U6  - https://doi.org/10.1002/celc.201801422
SN  - 2196-0216
VL  - 6
IS  - 6
SP  - 1732
EP  - 1737
PB  - Wiley-VCH
CY  - Weinheim
ER  - 
TY  - JOUR
A1  - Kaufmann, Hans Paul
A1  - Duffus, Benjamin R.
A1  - Mitrova, Biljana
A1  - Iobbi-Nivol, Chantal
A1  - Teutloff, Christian
A1  - Nimtz, Manfred
A1  - Jaensch, Lothar
A1  - Wollenberger, Ulla
A1  - Leimkühler, Silke
T1  - Modulating the Molybdenum Coordination Sphere of Escherichia coli Trimethylamie N-Oxide Reductase
JF  - Biochemistry
N2  - 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.
Y1  - 2018
U6  - https://doi.org/10.1021/acs.biochem.7b01108
SN  - 0006-2960
VL  - 57
IS  - 7
SP  - 1130
EP  - 1143
PB  - American Chemical Society
CY  - Washington
ER  -