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.
Deddish
P A
PA
Laboratory of Peptide Research, University of Illinois, Chicago College of Medicine 60612.
Skidgel
R A
RA
Erdös
E G
EG
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