Cloning, Sequence Analysis, and Distribution of Rat Metallocarboxypeptidase z

Abstract

A cDNA encoding human carboxypeptidase Z (CPZ), a novel metallocarboxypeptidase, was recently cloned (Song and Fricker, J. Biol. Chem., 272, 1054, 1997). In the present study, a cDNA encoding the rat homolog of CPZ was identified. As with the human form, rat CPZ contains an N-terminal domain of 120 amino acids that has 20% to 30% amino acid identity with the "frizzled" domain found on proteins that interact with Wnt, a protein involved in tissue polarity in early embryogenesis. Sequence analysis showed rat and human CPZ to be highly conserved within the frizzled domain (77% amino acid identity), the carboxypeptidase domain (91%), and the C-terminal 28 residues (78%). The entire rat CPZ protein has high sequence similarity with human CPZ (81% amino acid identity), moderate sequence similarity to human carboxypeptidase N (45%), human carboxypeptidase E (41%), and human carboxypeptidase M (33%), and less sequence similarity with other metallocarboxypeptidases. Northern blot analysis showed rat CPZ mRNA to be abundant in the placenta, with low to moderate levels in the brain, lung, thymus, and kidney. The BRL3A rat liver cell line and the PC12 rat adrenal cell line express high levels of CPZ mRNA. In situ hybridization analysis indicated that CPZ is expressed only in specific cell types. For example, in the brain, CPZ mRNA is present in leptomeningeal cells, but not in the majority of other cell types. This distribution in leptomeningeal cells is shared by AEBP1, a recently reported member of the metallocarboxypeptidase gene family. However, the distribution of CPZ and AEBP1 differ in pituitary and thyroid. Taken together, these studies suggest that CPZ functions in a range of cell types.

Head Bobber: An Insertional Mutation Causes Inner Ear Defects, Hyperactive Circling, and Deafness

“…Cpmx2 is a member of the metallocarboxypeptidase A family of digestive enzymes that are synthesized as inactive molecules with inhibitory propeptides that are cleaved to activate the enzyme. However, Cpxm2 lacks several of the predicted active site residues required for enzyme activity and may function instead as a phospholipid-binding protein (Xin et al 1998a(Xin et al , b, 1997. The Chst15 gene has highest sequence identity to human and rat type II transmembrane N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase (GalNAc4S6ST) found in the Golgi system required for biogenesis of glycoproteins (Ohtake et al 2001).…”

Characterization of the Enzymatic Properties of the First and Second Domains of Metallocarboxypeptidase D

“…Of these proteins, only CPD and carboxypeptidase Z demonstrate activity toward standard CPE substrates (27,(32)(33)(34). The cellular distribution of carboxypeptidase Z is restricted to specific cell types, such as the leptomeningeal cells in brain (36). In contrast, CPD has a broad tissue distribution and is present in many cell types in each tissue (37)(38)(39)(40).…”

Carboxypeptidase Z Is Present in the Regulated Secretory Pathway and Extracellular Matrix in Cultured Cells and in Human Tissues

“…One fusion protein contained the C-terminal 73 amino acids of rat CPZ. This region has 62% amino acid identity between rat and human CPZ, although the C-terminal 32 residues of this region has 81% amino acid identity (6,13). Another fusion protein contained 50 amino acids of rat CPZ corresponding to positions 163-212 (13).…”

“…However, based on the non-neuroendocrine tissue distribution of CPZ, it is unlikely that this enzyme plays a role in neuroendocrine peptide processing. For example, in brain, CPZ is primarily expressed in the leptomeningeal cells and is not detectable in neurons (13). Furthermore, the enzymatic properties of CPZ are not consistent with a role for this enzyme in the processing of peptides within the secretory pathway (6,14).…”

“…Thus, it is likely that CPZ performs another role, possibly outside the cell. CPZ is predicted to be secreted from cells based on the presence of an N-terminal signal sequence (6,13). When expressed in insect Sf9 cells using the baculovirus system, CPZ was not detected in the medium, although enzyme activity was detected on the cell surface despite the absence of a conventional membrane-binding domain (6).…”