LD-carboxypeptidase activity in Escherichia coli. II. Isolation, purification and characterization of the enzyme from E. coli K 12.

A LD-carboxypeptidase from Escherichia coli K 12 was isolated by Tris-EDTA treatment and purified to electrophoretic homogeneity by DEAE-cellulose chromatography. The enzyme has a molecular weight of approximately 12,000 as determined by sodium dodecyl sulfate-polyacrylamide electrophoresis and by Sephadex G-100 gel filtration. The studies of the substrate specificity of the enzyme revealed that UDP-MurNAc-tetrapeptide is a superior substrate, with a Km value of 1 X 10(-4) mol/l. The activity of the LD-carboxypeptidase was inhibited by D-amino acids and the beta-lactam antibiotic nocardicin A. Ki values of 0.3 and 43 mmol/l were determined for nocardicin A and D-homoserine, respectively. The properties of the purified enzyme correspond to activity I in ether treated cells.