N-Carbamyl-<scp>L</scp>-Amino Acid Amidohydrolase ofPseudomonassp. Strain NS671: Purification and Some Properties of the Enzyme Expressed inEscherichia coli

An N-carbamyl-L-amino acid amidohydrolase was purified from cells of Escherichia coli in which the gene for N-carbamyl-L-amino acid amidohydrolase of Pseudomonas sp. strain NS671 was expressed. The purified enzyme was homogeneous by the criterion of SDS-polyacrylamide gel electrophoresis. The results of gel filtration chromatography and SDS-polyacrylamide gel electrophoresis suggested that the enzyme was a dimeric protein with 45-kDa identical subunits. The enzyme required Mn2+ ion (above 1 mM) for the activity. The optimal pH and temperature were 7.5 and around 40 degrees C, respectively, with N-carbamyl-L-methionine as the substrate. The enzyme activity was inhibited by ATP and was lost completely with p-chloromercuribenzoate (1 mM). The enzyme was strictly L-specific and showed a broad substrate specificity for N-carbamyl-l-alpha-amino acids.