Evidence for pepstatin-sensitive conversion of porcine big endothelin-1 to endothelin-1 by the endothelial cell extract.
We designed a radioimmunoassay (RIA) specific for the C-terminal fragment (CTF, 22-39) of porcine big endothelin-1 (big ET-1, 1-39), and investigated whether the generation of ET-1 (1-21) is concomitant with that of the CTF during incubation of big ET-1 with the endothelial cell (EC)-extract. When the incubation mixture was applied to reverse-phase high performance liquid chromatography coupled with the RIAs for CTF and ET, immunoreactive (IR)-CTF eluted as one major and one minor peak, and IR-ET as one major peak. The retention times of each peak corresponded to those of synthetic CTF, big ET-1 and ET-1, respectively. An aspartic protease inhibitor pepstatin-A completely inhibited the generation of CTF- and ET-1-like materials. Both materials were also detected in culture medium of porcine aortic ECs. These findings strongly suggest that ET-1 is generated from big ET-1 via a single cleavage between Trp21 and Val22 in vascular ECs and that a pepstatin-sensitive aspartic protease is a possible candidate for big ET-1 converting enzyme.
Ikegawa
R
R
Department of Pharmacology, Osaka University of Pharmaceutical Sciences, Japan.
Matsumura
Y
Y
Takaoka
M
M
Morimoto
S
S
D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D016232:Endothelins; D004730:Endothelium, Vascular; D007700:Kinetics; D008970:Molecular Weight; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology*; D010436:Pepstatins / Q000494:pharmacology*; D010455:Peptides / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D013552:Swine
D016232:Endothelins; D009842:Oligopeptides; D010436:Pepstatins; D010455:Peptides