Purification and characterization of D-serine deaminase activator protein.
We purified the dsdC gene product, the specific activator of dsdA (D-serine deaminase) gene expression, to about 25% homogeneity from a strain in which its expression was amplified 100-fold. The purification involved, successively: DNase and high-salt treatment of cell extracts, DNA-cellulose chromatography, and Dyematrex (Amicon Corp.) column chromatography. We identified the protein as a discrete spot on two-dimensional O'Farrell gels after the DNA-cellulose step and quantitated it by densitometry. The active form was found to be a dimer. We estimated that there were eight activator dimers per wild-type cell. The activator is a slightly basic protein, with an experimental Km for its ligand D-serine of about 7 X 10(-6)M. The low concentration of the activator in wild-type cells and its autorepression may explain the previously observed partial dominance of dsdC+ in dsdCc/dsdC+ merodiploids.
D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005838:Genotype; D007700:Kinetics; D012695:L-Serine Dehydratase / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D013045:Species Specificity
D001426:Bacterial Proteins; D004274:DNA, Recombinant; D004262:DNA Restriction Enzymes; D012695:L-Serine Dehydratase