title: Kinetic regulation of a corrinoid-reducing metallo-ATPase by its substrates.
abstract: Corrinoid cofactors play a crucial role as methyl group carriers in the C1 metabolism of anaerobes, e.g. in the cleavage of phenyl methyl ethers by O-demethylases. For the methylation, the protein-bound corrinoid has to be in the super-reduced [Co(I) ]-state, which is highly sensitive to autoxidation. The reduction of inadvertently oxidized corrinoids ([Co(II) ]-state) is catalysed in an ATP-dependent reaction by RACE proteins, the reductive activators of corrinoid-dependent enzymes. In this study, a reductive activator of O-demethylase corrinoid proteins was characterized with respect to its ATPase and corrinoid reduction activity. The reduction of the corrinoid cofactor was dependent on the presence of potassium or ammonium ions. In the absence of the corrinoid protein, a basal slow ATP hydrolysis was observed which was obviously not coupled to corrinoid reduction. ATP hydrolysis was significantly stimulated by the corrinoid protein in the [Co(II) ]-state of the corrinoid cofactor. The stoichiometry of ATP hydrolysed per mol corrinoid reduced was near 1:1. Site-directed mutagenesis was applied to study the impact of a highly conserved region possibly involved in nucleotide binding of RACE proteins, indicating that an aspartate and a glycine residue may play an essential role for the function of the enzyme.
authors.affiliation: Institut für Mikrobiologie, Friedrich-Schiller-Universität Jena, Lehrstuhl für Angewandte und Ökologische Mikrobiologie, Philosophenweg 12, 07743, Jena, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Sperfeld
authors.forename: Gabriele
authors.initials: G
authors.lastname: Diekert
authors.forename: Sandra
authors.initials: S
authors.lastname: Studenik
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D064751:Ammonium Compounds / Q000378:metabolism; D003067:Coenzymes; D045728:Corrinoids / Q000378:metabolism*; D004252:DNA Mutational Analysis; D006868:Hydrolysis; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D011188:Potassium / Q000378:metabolism
chemical_list: D064751:Ammonium Compounds; D003067:Coenzymes; D045728:Corrinoids; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D011188:Potassium