Purification and kinetic properties of a D-amino-acid peptide hydrolyzing enzyme from pig kidney cortex and its tentative identification with renal membrane dipeptidase.

We previously reported the presence of an enzyme activity which hydrolyzes Gly-D-Asp in pig kidney cortex. In the present study, an enzyme which hydrolyzes this peptide and other peptides having a low number of D- and L-amino acids has been purified from the brush border membranes of the same tissue. The native enzyme, having a molecular weight of 99,000, was apparently a homodimer of a subunit with a molecular weight of 48,000 and its optimum pH was 7.8 with Gly-D-Ala as substrate. The enzyme hydrolyzed many dipeptides, but not most of the tripeptides tested with a few exceptions, from which the carboxyl-terminal amino acid was liberated by the enzyme. Of the dipeptides examined, those having a D-amino acid at the amino-terminal were poor substrates, whereas those bearing a D-amino acid at the carboxyl-terminal were good substrates, comparable with their diastereomers with a L-amino acid at the same position. The enzyme was potently inhibited by cilastatin but not by amastatin and bestatin. Metal ion chelators and dithiothreitol were also inhibitory. Comparison of the properties of present enzyme with those of other known enzymes suggests that this should be tentatively identified with renal membrane dipeptidase. The demonstrated high activity toward dipeptides containing various D-amino acids at the carboxyl-terminal suggests a possibility that the enzyme in fact plays a role in degradation in vivo of D-amino-acid-containing peptides.