Enhanced Co2+ activation and inhibitor binding of carboxypeptidase M at low pH. Similarity to carboxypeptidase H (enkephalin convertase).
Carboxypeptidases H and M differ in their distribution and other properties, but both are activated by Co2+ and inhibited by guanidinoethylmercaptosuccinic acid. The higher degree of activation or inhibition of carboxypeptidase H by these agents at acid pH has been employed to identify this enzyme in tissues. We found that the activation or inhibition of both purified and plasma-membrane-bound human carboxy-peptidase M depends on the pH of the medium. CoCl2 activated over 6-fold at pH 5.5, but less than 2-fold at pH 7.5. Guanidinoethylmercaptosuccinic acid inhibited the membrane-bound carboxypeptidase M more effectively than the purified enzyme, and the IC50 was about 25-30 times lower at pH 5.5. As purified human plasma carboxypeptidase N and pancreatic carboxypeptidase B were also activated more at pH 5.5, we conclude that the increased activation by CoCl2 is due to the enhanced dissociation of Zn2+ below the pKa of the ligands that co-ordinate the cofactor in the protein. Thus increased activation or inhibition at acid pH would not differentiate basic carboxypeptidases.
Laboratory of Peptide Research, University of Illinois, Chicago College of Medicine 60612.
P A
PA
Deddish
R A
RA
Skidgel
E G
EG
Erdös
D003035:Cobalt / Q000494:pharmacology*; D004789:Enzyme Activation; D058851:GPI-Linked Proteins; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008871:Microvilli / Q000201:enzymology; D013386:Succinates / Q000494:pharmacology*
D058851:GPI-Linked Proteins; D013386:Succinates; D003035:Cobalt; C022014:guanidinoethylmercaptosuccinic acid; C058479:carboxypeptidase M; D008666:Metalloendopeptidases; C018021:cobaltous chloride