The search result changed since you submitted your search request. Documents might be displayed in a different sort order.
  • search hit 5 of 10
Back to Result List

Identification of a bis-molybdopterin intermediate in molybdenum cofactor biosynthesis in escherichia coli

  • The molybdenum cofactor is an important cofactor, and its biosynthesis is essential for many organisms, including humans. Its basic form comprises a single molybdopterin (MPT) unit, which binds a molybdenum ion bearing three oxygen ligands via a dithiolene function, thus forming Mo-MPT. In bacteria, this form is modified to form the bis-MPT guanine dinucleotide cofactor with two MPT units coordinated at one molybdenum atom, which additionally contains GMPs bound to the terminal phosphate group of the MPTs (bis-MGD). The MobA protein catalyzes the nucleotide addition to MPT, but the mechanism of the biosynthesis of the bis-MGD cofactor has remained enigmatic. We have established an in vitro system for studying bis-MGD assembly using purified compounds. Quantification of the MPT/molybdenum and molybdenum/phosphorus ratios, time-dependent assays for MPT and MGD detection, and determination of the numbers and lengths of Mo-S and Mo-O bonds by X-ray absorption spectroscopy enabled identification of a novel bis-Mo-MPT intermediate on MobAThe molybdenum cofactor is an important cofactor, and its biosynthesis is essential for many organisms, including humans. Its basic form comprises a single molybdopterin (MPT) unit, which binds a molybdenum ion bearing three oxygen ligands via a dithiolene function, thus forming Mo-MPT. In bacteria, this form is modified to form the bis-MPT guanine dinucleotide cofactor with two MPT units coordinated at one molybdenum atom, which additionally contains GMPs bound to the terminal phosphate group of the MPTs (bis-MGD). The MobA protein catalyzes the nucleotide addition to MPT, but the mechanism of the biosynthesis of the bis-MGD cofactor has remained enigmatic. We have established an in vitro system for studying bis-MGD assembly using purified compounds. Quantification of the MPT/molybdenum and molybdenum/phosphorus ratios, time-dependent assays for MPT and MGD detection, and determination of the numbers and lengths of Mo-S and Mo-O bonds by X-ray absorption spectroscopy enabled identification of a novel bis-Mo-MPT intermediate on MobA prior to nucleotide attachment. The addition of Mg-GTP to MobA loaded with bis-Mo-MPT resulted in formation and release of the final bis-MGD product. This cofactor was fully functional and reconstituted the catalytic activity of apo-TMAO reductase (TorA). We propose a reaction sequence for bis-MGD formation, which involves 1) the formation of bis-Mo-MPT, 2) the addition of two GMP units to form bis-MGD on MobA, and 3) the release and transfer of the mature cofactor to the target protein TorA, in a reaction that is supported by the specific chaperone TorD, resulting in an active molybdoenzyme.show moreshow less

Export metadata

Additional Services

Search Google Scholar Statistics
Metadaten
Author details:Stefan ReschkeGND, Kajsa G. V. Sigfridsson, Paul Kaufmann, Nils Leidel, Sebastian Horn, Klaus Gast, Carola Schulzke, Michael HaumannORCiD, Silke LeimkühlerORCiDGND
DOI:https://doi.org/10.1074/jbc.M113.497453
ISSN:0021-9258
ISSN:1083-351X
Title of parent work (English):The journal of biological chemistry
Publisher:American Society for Biochemistry and Molecular Biology
Place of publishing:Bethesda
Publication type:Article
Language:English
Year of first publication:2013
Publication year:2013
Release date:2017/03/26
Volume:288
Issue:41
Number of pages:10
First page:29736
Last Page:29745
Funding institution:Deutsche Forschungsgemeinschaft (DFG) Cluster of Excellence "Unifying Concepts in Catalysis"; DFG [LE1171/6-1, Ha3265/3-1, Ha3265/6-1]; Heisenberg Fellowship; European Cooperation in Science and Technology (COST) Action [CM1003]; Stiftelsen Bengt Lundqvist minne; Wenner-Gren Foundation; European Research Council (ERC)
Organizational units:Mathematisch-Naturwissenschaftliche Fakultät / Institut für Biochemie und Biologie
Peer review:Referiert
Accept ✔
This website uses technically necessary session cookies. By continuing to use the website, you agree to this. You can find our privacy policy here.