CLONING AND SEQUENCE-ANALYSIS OF CDNA-ENCODING RAT CARBOXYPEPTIDASE-D

Carboxypeptidase D (CPD) is a recently described 180-kD enzyme with ca rboxypeptidase E-like enzymatic properties, CPD has been proposed to b e present in the secretory pathway and to contribute to peptide hormon e processing in the Cpe(fat)/Cpe(fat) mouse, which lacks functional CP E. Sequence analysis of cDNA clones encoding rat CPD show the protein to contain an amino-terminal signal peptide, three carboxypeptidase-li ke domains, a putative transmembrane domain, and a 60-amino-acid cytop lasmic tail, Whereas active site, substrate-binding, and metal-binding residues of other metallocarboxypeptidases are conserved in the first two domains of CPD, several of the critical residues are not conserve d in the third domain; this third domain is not predicted to form an a ctive carboxypeptidase, The overall homology between rat CPD and the d uck homolog gp180 is high, with 75% amino acid identity, The three car boxypeptidase domains show 66%, 83%, and 82% amino acid identity betwe en rat CPD and duck gp180, Homology is also high in the transmembrane domain (86%) and in the cytoplasmic tail (97%), The mouse Cpd gene map s to the medial portion of chromosome 11, approximately 45.5 cM distal to the centromere, Northern blot analysis of CPD mRNA shows major ban ds of approximately 8 and 4 kb in many rat tissues,and additional spec ies ranging from 1.4 to 5 kb that are expressed in some tissues or cel l lines, CPD mRNA is detectable in most tissues examined, and is most abundant in hippocampus, spinal cord, atrium of the heart, colon, test is, and ovaries, In situ hybridization of CPD mRNA shows a distributio n in many cells in rat brain and other tissues, with high levels in hi ppocampus, olfactory bulb, and the intermediate pituitary, The broad d istribution is consistent with a role for CPD in the processing of man y peptides and proteins that transit the secretory pathway.