Cloning and expression of human carboxypeptidase Z, a novel metallocarboxypeptidase.
A novel cDNA, designated carboxypeptidase Z (CPZ), was identified based on its homology to known metallocarboxypeptidases. Northern blot analysis shows bands of 2.1 and/or 2.6 kilobases in all tissues examined. The major form of CPZ mRNA in human salivary gland encodes a protein with an open reading frame of 641 amino acids. In addition, three variants were found that presumably arise due to alternative intron splicing. The 641-amino acid protein contains an 18-residue signal peptide-like sequence, a 120-residue region that shows 23-29% amino acid identity with a Cys-rich domain found in frizzled proteins and in type XVIII collagen, and then a 390-residue carboxypeptidase domain with 49% amino acid identity to carboxypeptidases E and N. The 641-amino acid form of CPZ expressed in the baculovirus system cleaves 5-dimethylaminonaphthalene-1-sulfonyl (dansyl)-Phe-Ala-Arg, although the level of enzyme activity was approximately 10-fold lower than either carboxypeptidase E or D expressed using the same viral system. The CPZ activity is more active at neutral pH than at pH 5.5 and is inhibited by active site-directed inhibitors of metallocarboxypeptidases. In summary, CPZ is a novel metallocarboxypeptidase that is active toward substrates with C-terminal basic amino acids.
Song
L
L
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Fricker
L D
LD
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002268:Carboxypeptidases / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary; D006801:Humans; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D012469:Salivary Glands / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid
D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins; D002268:Carboxypeptidases; C118337:metallocarboxypeptidase Z