A NOVEL PROTEINASE, GLUCAGON-DEGRADING ENZYME, SECRETED BY A HUMAN PANCREATIC-CANCER CELL-LINE, HPC-YO

Sixty-four kinds of cell lines were examined as to their ability to de grade glucagon using conditioned-media obtained from their protein-fre e cultures. Two human tumor cell lines were shown to produce this acti vity, and the cell line, HPC-YO, established from a human pancreatic c arcinoma was shown to produce the highest level of activity. The gluca gon-degrading enzyme (GDE) was purified from HPC-YO conditioned-medium by a combination of ion-exchange, gel filtration, and hydroxylapatite column chromatographies. The purified GDE also degraded vasoactive in testinal polypeptide (VIP) and secretin, however, it did not cleave EG F, gastrin, insulin, somatostatin, substance P, neurotensin, or growth hormone. The molecular weight of GDE is 83,000, as determined on SDS- polyacrylamide gel electrophoresis. The N-terminal amino acid sequence of GDE was blocked, and the five partial amino acid sequences obtaine d on lysyl-endopeptidase digestion were determined to be N-L-T-E-E-Y-D -V-S-D-G-E-I-E-L-L-Y-E-K, V-E-T-Y-Y-D-L-L-F-E-K, L-Y-W-F-L-D-E-A-K, S- N-S-T-S-Y-V-K, and Y-Y-A-S-T-S-Y-D-D-T-Y-K. The same or homologous ami no acid sequences have not been found in known proteins, demonstrating that GDE is a novel peptidase that degrades the secretin family: gluc agon, VIP, and secretin.