Cloning and functional expression of endothelin-converting enzyme from rat endothelial cells.
Endothelin (ET) is a 21-residue potent vasoconstrictive peptide produced by vascular endothelial cells and formed from its precursor, big endothelin (big ET), by endothelin-converting enzyme (ECE). Here we report the cloning and functional expression of a complementary DNA encoding a rat ECE from endothelial cells. Rat ECE is a highly glycosylated protein consisting of 10 possible N-linked glycosylation sites, a zinc-binding domain, and a single membrane-spanning region. It has structural and sequence homology to neutral endopeptidase and Kell blood group protein, a putative neutral endopeptidase. Monoclonal antibody risen against purified rat lung ECE recognized broad glycosylated bands in membrane fractions prepared from both rat lung and COS cells transfected with a rat ECE expression vector. Expressed ECE was inhibited by phosphoramidon, but not by thiorphan. It also cleaved big ET-1 efficiently but not big ET-2 or big ET-3. Northern blot analysis revealed that ECE messenger RNA is widely expressed in many tissues.
Shimada
K
K
Biological Research Laboratories, Sankyo Co., Ltd., Tokyo, Japan.
Takahashi
M
M
Tanzawa
K
K
D000595:Amino Acid Sequence; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000096:biosynthesis* / Q000378:metabolism; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular / Q000379:methods; D016384:Consensus Sequence; D017931:DNA Primers; D018076:DNA, Complementary / Q000378:metabolism; D000072396:Endothelin-Converting Enzymes; D004730:Endothelium, Vascular / Q000201:enzymology*; D015870:Gene Expression; D006801:Humans; D007700:Kinetics; D008168:Lung / Q000201:enzymology; D008666:Metalloendopeptidases; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; D000072396:Endothelin-Converting Enzymes