251 – Metallocarboxypeptidase D

Publisher Summary This chapter examines the activity, specificity and structural chemistry of metallocarboxypeptidase D. It is found that when the cDNA encoding gpl80 was cloned and sequenced, it became apparent that this protein represented a 180 kDa member of the metallocarboxypeptidase family. Carboxypeptidase D (CPD) removes C-terminal +Lys or + Arg residues from a variety of substrates, with no detectable activity towards nonbasic residues. It prefers an Ala in the penultimate position. In comparing N-terminally blocked small peptides, CPD shows lower activity with dipeptides than with tripeptides. The optimal pH for CPD activity is 5–7. CPD is stimulated by millimolar concentrations of Co2+ and inhibited by 1,10-phenanthroline. The enzyme is partially inhibited by some thiol-directed reagents, such as 7-chloromercuriphenylsulfonate or HgCl2. CPD is a single-chain protein that exists in several forms, with the major form being 180 kDa. It is likely that CPD contains 2–3 mol of zinc per mol of protein based on the homology between CPD and other metallocarboxypeptidases. CPD contains three carboxypeptidase domains of about 390 amino acids, which are followed by a transmembrane domain and a short 60 residue sequence that forms the cytosolic tail.