YAMAMURA Yoshiro Koyto Prefectural University of Medicine, Department of Cell Biology, Instructor, 医学部, 助手 (10220443)
IMANISHI Jiro Kyoto Prefectural University of Medicine, Department of Microbiology, Professor, 医学部, 教授 (40112510)
|Budget Amount *help
¥2,000,000 (Direct Cost : ¥2,000,000)
Fiscal Year 1993 : ¥2,000,000 (Direct Cost : ¥2,000,000)
We have found a novel proteinase which degrades endothelin. We designated this proteinase as endothelin-degrading enzyme (EDE) at first. This enzyme also cleaves other peptide hormones such as glucagon, vasoactive intestinal peptide (VIP) and secretin, however, it did not cleave EGF,gastrin, insulin, somatostatin, substance P,neurotensin, or growth hormone. We renamed this enzyme as glucagon-degrading enzyme (GDE), since the affinty to glucagon was higher than endothelin. This was purified from protein-free cultured human pancreatic cancer cell line, HPC-YO conditioned-medium by a combination of ion-exchange, gel filtration and hydroxlapatite column chromatographies. The molecular weight of GDE is 83,000, as determined on SDS-PAGE.The N-terminal amino acid sequence of GDE was blocked, and the five partial amino acid sequences obtained onlysyl-endopeptidase digestion were determined to be NLTEEYDVS-DGELELLYEK,VETYYDLLFEK,LYWFLDEAK,SNSTSYVK,and YYASTSYDDTYK.The same or homologous amino acid sequences have not been found in known proteins, demonstrating that GDE is a novel peptidase. Furthermore, We could clarified open reading frame (ORF) of GDE cDNA,and the same or homologous DNA sequences have not found. The ORF was consisted of 2.6 kbp. We produced several kinds of antibodies to GDE,and established the microassay system of GDE in the conditioned media of various kind of cell lines. In near future, we will measure GDE levels in sera of healthy controls and patitents.