Some physical and kinetic properties of adenylyl sulfate reductase from Desulfovibrio vulgaris.
Adenylyl sulfate reductase has been purified from the anaerobic sulfate-reducing bacterium, Desulfovibrio vulgaris, and judged to be homogenous by several criteria. Different forms of the enzyme could be visualized in polyacrylamide gels after electrophoresis and these polymeric species have been studied by a combination of absorption spectra, polyacrylamide gel electrophoresis, and sedimentation  velocity experiments. A dimeric species of molecular weight 440,000 is stable in potassium phosphate buffer but can be dissociated to a 220,000 molecular weight species by either changing the buffer system to Tris-maleate or addition of AMP, DAMP, or adenylyl sulfate. Other catalytically active nucleotides are not capable of effecting this dissociation. The enzyme was determined to contain 12 non-heme irons, 12 acid-labile sulfides, and 1 FAD per molecule when calculated on the basis of a monomeric molecular weight of 220,000. ;el electrophoresis in the presence of sodium dodecyl sulfate indicated subunits of molecular weight 72,000 and 20,000. The extinction coefficient when determined in phosphate buffer at 372 nm is 108,000 M-1 cm-a. Steady state kinetic experiments employing ferricyanide, cytochrome c, and reduced methyl viologen as artificial electron transfer reagents were performed and the kinetic constants obtained under various conditions. Several nucleotide substrates were employed and compared in each assay with respect to Km and Vmax. The reduction of cytochrome c was found to be sensitive to both anaerobiosis and superoxide dismutase, suggesting the involvement of superoxide anions with this electron acceptor.
D000249:Adenosine Monophosphate; D000535:Aluminum; D002845:Chromatography; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D003574:Cytochrome c Group; D003901:Desulfovibrio / Q000201:enzymology*; D005292:Ferricyanides; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D010269:Paraquat; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013431:Sulfates
D003574:Cytochrome c Group; D005292:Ferricyanides; D046911:Macromolecular Substances; D013431:Sulfates; D000249:Adenosine Monophosphate; D000535:Aluminum; D010088:Oxidoreductases; D010269:Paraquat