PEPTIDASE-D OF ESCHERICHIA-COLI-K-12, A METALLOPEPTIDASE OF LOW SUBSTRATE-SPECIFICITY

Peptidase D of Escherichia coli was overproduced from a multicopy plas mid and purified to electrophoretic homogeneity. The pure enzyme was s table at 4 degrees C or - 20 degrees C and had a pH optimum at pH 9, a nd a pI of 4.7; the temperature optimum was at 37 degrees C. As the en zyme was activated by Co2+ and Zn2+, and deactivated by metal chelator s, it appears to be a metallopeptidase. By activity staining of native gels, 11 dipeptides which are preferentially cleaved by peptidase D w ere identified. Peptidase D activity required dipeptide substrates wit h an unblocked amino terminus and the amino group in the cu or beta po sition. Non-protein amino acids and proline were not accepted in the C -terminal position, whereas some dipeptide amides and formyl amino aci ds were hydrolyzed. K-m values of 2 to 5 mM indicate a relatively poor interaction of the enzyme with its substrates.