Characterization of endothelin receptors ETA and ETB expressed in COS cells.
The cloned cDNA genes for endothelin receptors ETA and ETB were expressed in COS cells, and the binding characteristics of the two receptors with three isopeptide ligands (ET-1, ET-2, and ET-3) were examined in detail. The results indicated that the stability of receptor-ET-1 complexes formed with ETA and ETB were significantly different from each other, while their affinities to ET-1 were similar. The preformed ETA-ET-1 complex readily dissociated upon SDS-PAGE, as did many of the other receptors so far studied, while the ETB-ET-1 complex survived SDS-PAGE when it was run at low temperature (approximately 4 degrees C). Clear differences in stability were also shown in comparative studies of acid treatment of the two types of complexes. Only the ETB-ET-1 complex was resistant to acid treatment (0.2 M acetic acid, 0.5 M NaCl), and its 50 kDa monomeric receptor-ligand complex remained intact. The ETB-ET-1 complex (50 kDa) formed at 4 degrees C on the surface of COS cells, however, was susceptible to limited proteolysis at 37 degrees C that reduced the molecular size of the complex to a distinct 35 kDa. No such size reduction was observed with the preformed ETA-ET-1 complex. The overall structure of two endothelin receptors, as deduced from the sequence of cloned cDNAs, is similar in many respects. However, the present findings demonstrate distinct differences in the biochemical nature of the two receptors, which suggest their distinct biological functions.
Takasuka
T
T
Department of Molecular Genetics, Nippon Roche Research Center, Kanagawa.
Adachi
M
M
Miyamoto
C
C
Furuichi
Y
Y
Watanabe
T
T
D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D004355:Drug Stability; D016232:Endothelins / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D017466:Receptors, Endothelin / Q000235:genetics* / Q000378:metabolism
D016232:Endothelins; D017466:Receptors, Endothelin; D004247:DNA