Tissue distribution and characterization of soluble and membrane-bound forms of metallocarboxypeptidase D.
Metallocarboxypeptidase D (CPD) is a recently discovered 180-kDa membrane-bound carboxypeptidase E-like enzyme (Song, L. and Fricker, L. D. (1995) J. Biol. Chem. 270, 25007-25013). In the present study, a soluble CPD-like activity has been purified to homogeneity and characterized. On denaturing polyacrylamide gels, the soluble enzyme from bovine pituitary glands appears as two bands of 170 and 135 kDa which are converted to 155 and 115 kDa by endoglycosidase F. Both of the soluble forms of CPD are recognized by an antisera raised against CPD purified from rat brain membranes. The partial N-terminal amino acid sequences of the two soluble forms are identical to each other and to the predicted N terminus of duck gp180. The soluble and membrane forms of CPD have similar pH optima, inhibitor specificities, and kinetic parameters for substrate hydrolysis. CPD-like enzymatic activity is detected in all rat tissues examined, with highest levels in pituitary, brain, and adrenal. Western blot analysis indicates that both soluble and membrane forms of CPD are present in rat brain, heart, liver, and kidney. At least four distinct 100-180-kDa forms of CPD are detected on Western blots, although an antiserum raised against the C-terminal region of rat CPD recognizes only the 180-kDa membrane-bound form. The finding that CPD is widely distributed suggests a broad role for this enzyme in the processing of proteins that transit the secretory pathway.
Song
L
L
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA. fricker@aecom.yu.edu
Fricker
L D
LD
D000818:Animals; D002268:Carboxypeptidases / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D010902:Pituitary Gland / Q000201:enzymology; D051381:Rats; D012995:Solubility; D013379:Substrate Specificity
D002268:Carboxypeptidases; C078874:carboxypeptidase D