THE SEQUENCE AND CONFORMATION OF HUMAN PANCREATIC PROCARBOXYPEPTIDASEA2 - CDNA CLONING, SEQUENCE-ANALYSIS, AND 3-DIMENSIONAL MODEL

A full-length cDNA clone coding for human pancreatic preprocarboxypept idase A2 has been isolated from a lambda gt11 human pancreatic library , Expression clones were identified by specific interaction with antis era raised against the native protein. The open reading frame of the p olynucleotide sequence is 1254 base pairs in length and encodes a prot ein of 417 amino acids, This cDNA includes a short leader signal pepti de of 16 amino acids and a 94-amino acid-long activation segment, The amino acid sequence shows 89% identity to that of rat procarboxypeptid ase A2, the only A2 form sequenced so far, and 64% identity to that of human procarboxypeptidase A1. The newly determined sequence was model ed to the three-dimensional crystal structures of both bovine carboxyp eptidase A and porcine procarboxypeptidase A1 by a novel distance geom etry approach, Biases in the modeling were avoided by relying exclusiv ely on automatic procedures and by using random structures as starting points, Information taken from the known homologous structures refers only to the backbone since no explicit data describing the conformati on of side chains were transferred, Ten structures of human carboxypep tidase A2 were determined on the basis of each of the two known crysta l structures, The root-mean-square distance for the backbone atoms bet ween the 10 structures and their mean for 237 selected residues is 0.7 Angstrom when starting from the bovine protein and 0.8 Angstrom for 2 51 selected residues when starting from the porcine protein. The 94 re sidue-long activation segment was also determined in the modeling base d on the porcine zymogen; its structure is well defined but not its or ientation with respect to the enzyme moiety. The model obtained for hu man procarboxypeptidase A2 is discussed with respect to the specificit y and activation of the enzyme.