PMID:11507093

Citation	Jeon, WB, Cheng, J and Ludden, PW (2001) Purification and characterization of membrane-associated CooC protein and its functional role in the insertion of nickel into carbon monoxide dehydrogenase from Rhodospirillum rubrum. J. Biol. Chem. 276:38602-9
Abstract	The accessory protein CooC, which contains a nucleotide-binding domain (P-loop) near the N terminus, participates in the maturation of the nickel center of carbon monoxide dehydrogenase (CODH). In this study, CooC was purified from the chromatophore membranes of Rhodospirillum rubrum with a 3,464-fold purification and a 0.8% recovery, and its biochemical properties were characterized. CooC is a homodimer with a molecular mass of 61-63 kDa, contains less than 0.1 atom of Ni(2+) or Fe(2+) per dimer, and has a lambda(max) at 277.5 nm (epsilon(277.5) 32.1 mm(-1) cm(-1)) with no absorption peaks at the visible region. CooC catalyzes the hydrolysis of ATP and GTP with K(m) values of 24.4 and 26.0 microm and V(max) values of 58.7 and 3.7 nmol/min/mg protein for ATP and GTP hydrolysis, respectively. The P-loop mutated form of K13Q CooC was generated by site-specific replacement of lysine by glutamine and was purified according to the protocol for wild-type CooC purification. The K13Q CooC was inactive both in ATP hydrolysis and in vivo nickel insertion. In vitro nickel activation of apoCODH in the cell extracts from UR2 (wild type) and UR871 (K13Q CooC) showed that activation of nickel-deficient CODH was enhanced by CooC and dependent upon ATP hydrolysis. The overall results suggest that CooC couples ATP hydrolysis with nickel insertion into apoCODH. On the basis of our results and models for analogous systems, the functional roles of CooC in nickel processing into the active site of CODH are presented.
Links	PubMed Online version:10.1074/jbc.M104945200
Keywords	Adenosine Triphosphatases/metabolism; Adenosine Triphosphate/metabolism; Aldehyde Oxidoreductases/chemistry; Aldehyde Oxidoreductases/metabolism; Bacterial Proteins/chemistry; Bacterial Proteins/isolation & purification; Bacterial Proteins/physiology; Binding Sites; Blotting, Western; Cell Membrane/metabolism; Dimerization; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Escherichia coli Proteins; GTP Phosphohydrolases/metabolism; Genotype; Hydrolysis; Kinetics; Membrane Proteins; Multienzyme Complexes/chemistry; Multienzyme Complexes/metabolism; Mutation; Nickel/chemistry; Nickel/metabolism; Protein Binding; Protein Structure, Tertiary; Rhodospirillum rubrum/metabolism; Spectrophotometry; Time Factors; Ultraviolet Rays
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
RHORU:COOC	part_of	GO:0016020: membrane	ECO:0000314: direct assay evidence used in manual assertion		C		Seeded From UniProt	complete
RHORU:COOC	enables	GO:0003924: GTPase activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
RHORU:COOC		GO:0018414: nickel incorporation into metallo-sulfur cluster	ECO:0000314: direct assay evidence used in manual assertion		P		Figure 5 shows both CooC and CooJ are required for insertion of the Ni-Iron center into CooS	complete CACAO 3360
RHORU:COOC		GO:0042623: ATPase activity, coupled	ECO:0000314: direct assay evidence used in manual assertion		F		Figure 3 shows the ATPase activity of CooC	complete CACAO 3362
RHORU:COOC		GO:0003924: GTPase activity	ECO:0000314: direct assay evidence used in manual assertion		F		Figure 3 shows that CooC has GTPase activity	complete CACAO 3363
RHORU:COOC		GO:0016020: membrane	ECO:0000314: direct assay evidence used in manual assertion		C		Figure 2	complete CACAO 3367
RHORU:COOC	enables	GO:0042623: ATPase activity, coupled	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
RHORU:P72321	Contributes to	GO:0018414: nickel incorporation into metallo-sulfur cluster	ECO:0000314:		P		Figure 5 shows that CooC and CooJ are required for insertion of the Ni-Iron center into CooS	complete CACAO 3359
RHORU:P72321		GO:0018414: nickel incorporation into metallo-sulfur cluster	ECO:0000314:		P		Figure 5	complete CACAO 3369

References

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