Human endothelin-converting enzyme (ECE-1): three isoforms with distinct subcellular localizations.
Endothelin-converting enzyme 1 (ECE-1) is a membrane-bound metalloprotease that catalyses the conversion of inactive big endothelins into active endothelins. Two different isoforms (ECE-1a and ECE-1b) have previously been identified for human ECE-1. In the present study we have cloned a novel human ECE-1 isoform, termed ECE-1c, and have thus shown for the first time the existence of three distinct ECE-1 isoforms. The three isoforms differ only in their N-terminal regions and are derived from a single gene through the use of alternative promoters. Ribonuclease protection experiments revealed that, although the relative levels of the three isoform mRNA species vary between human tissues, ECE-1c mRNA is generally the predominant isoform messenger. Immunofluorescence microscopy analysis showed distinct subcellular localizations for the three isoforms: whereas ECE-1a and ECE-1c are localized at the cell surface, ECE-1b was found to be intracellular and showed significant co-localization with a marker protein for the trans-Golgi network. We determined that the three isoforms have similar kinetic rate constants (Km, kcat and Vmax) for the processing of big endothelin 1 and that the big endothelin isoforms 1, 2 and 3 are cleaved with similar relative velocities of 1.0:0.1:0.1 by the three isoenzymes.
Schweizer
A
A
F. Hoffmann-La Roche Ltd., Pharma Division, Preclinical Research, Grenzacherstrasse 124, CH-4070 Basel, Switzerland.
Valdenaire
O
O
Nelböck
P
P
Deuschle
U
U
Dumas Milne Edwards
J B
JB
Stumpf
J G
JG
Löffler
B M
BM
D000595:Amino Acid Sequence; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000032:analysis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology*; D003001:Cloning, Molecular; D006224:Cricetinae; D019332:Endothelin-1; D000072396:Endothelin-Converting Enzymes; D016232:Endothelins / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000201:enzymology*; D006801:Humans; D007527:Isoenzymes / Q000032:analysis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008666:Metalloendopeptidases / Q000032:analysis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011498:Protein Precursors / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D017422:Sequence Analysis, DNA
D019332:Endothelin-1; D016232:Endothelins; D007527:Isoenzymes; D011498:Protein Precursors; D012333:RNA, Messenger; D012260:Ribonucleases; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; C000606923:ECE1 protein, human; D000072396:Endothelin-Converting Enzymes