Endothelin-2-converting enzyme from human renal adenocarcinoma cells is a phosphoramidon-sensitive, membrane-bound metalloprotease.
From the membrane fraction of cultured human renal adenocarcinoma (ACHN) cells, two endothelin-2-converting enzymes (ECE-2A and ECE-2B) were solubilized with detergent Lubrol PX and separated by hydrophobic butyl fast-performance liquid chromatography. The pH range of the converting activity of ECE-2B for big endothelin-1 (big ET-1), big ET-2, or big ET-3, was very narrow, and the optimal pH for each substrate was significantly different; the pH optimum for big ET-1 was 6.8 and that for big ET-2 or big ET-3 was 6.4. The ET-converting activity was abolished by phosphoramidon, 1,10-phenanthroline and EDTA but was not inhibited by thiorphan, E-64, leupeptin, PCMS, p-APMSF, or pepstatin A. The conversion efficiency for big ET-2 or big ET-3 by ECE-2B was approximately one-eighth of that for big ET-1. The molecular weight of ECE-2B was estimated to be 400 kDa by gel filtration. Because these characteristics of ECE-2B are very similar to those of ET-1-converting enzyme (ECE-1) in endothelial cells, these results raise the possibility that ECE-2B is identical to ECE-1 and that a single ECE physiologically converts all big ETs to the corresponding ETs in ET-producing cells.
Shinmi
O
O
Center for Biomedical Science, Chiba University, School of Medicine, Japan.
Yorimitsu
K
K
Moroi
K
K
Nishiyama
M
M
Sugita
Y
Y
Saito
T
T
Inagaki
Y
Y
Masaki
T
T
Kimura
S
S
D000230:Adenocarcinoma / Q000201:enzymology*; D016282:Aspartic Acid Endopeptidases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D000072396:Endothelin-Converting Enzymes; D016232:Endothelins / Q000378:metabolism*; D006020:Glycopeptides / Q000494:pharmacology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007680:Kidney Neoplasms / Q000201:enzymology*; D008666:Metalloendopeptidases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000494:pharmacology; D011863:Radioimmunoassay; D014407:Tumor Cells, Cultured
D016232:Endothelins; D006020:Glycopeptides; D011480:Protease Inhibitors; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; C000606923:ECE1 protein, human; D000072396:Endothelin-Converting Enzymes; C008890:phosphoramidon