Subunits of luteinizing hormone-human chorionic gonadotropin receptor from bovine corpora lutea.
A batch of 24 mg of luteinizing hormone-human chorionic gonadotropin (LH-hCG) receptor was isolated from bovine corpora lutea. The LH-hCG receptor showed specific binding with hCG. The receptor-hCG complex activated the regulatory Ns protein isolated from rabbit liver, which in turn stimulated adenylate cyclase to convert ATP into cAMP in vitro, attesting to the biological activity of the purified LH-hCG receptor. The LH-hCG receptor was treated with 2% sodium dodecyl sulfate (SDS) to prepare the molecular weight (Mr) 280K dimer and with 50 mM dithiothreitol (DTT) to prepare the Mr 120K monomer and subunits of Mr 85K and 38K. Oligomers of various molecular weights were recovered from gel filtration columns due to the reassociation of disulfide bonds between monomers and subunits. Hence, the receptor monomer was also dissociated into subunits of Mr 85K and 38K by reduction of -S-S-bonds with 50 mM DTT in 2% SDS and alkylation of sulfhydryl groups in the presence of 100 mM N-ethyl-maleimide. The subunits were separated by gel filtration through columns of Ultrogel AcA-44 and Sephadex G-75. The yields of the purified alkylated subunits of Mr 85K and 38K were 1.8 and 1.5 mg, respectively. Each subunit migrated as a single entity in SDS-polyacrylamide gel electrophoresis. The monomer of the receptor of Mr 120K showed specific binding with 125I-hCG, suggesting it to be the minimum molecular weight functional unit of the receptor. The Mr 85K and 38K subunits bound 125I-hCG, which could not be displaced with unlabeled hCG.(ABSTRACT TRUNCATED AT 250 WORDS)
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002417:Cattle; D006063:Chorionic Gonadotropin / Q000378:metabolism; D003338:Corpus Luteum / Q000378:metabolism*; D004789:Enzyme Activation; D005260:Female; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011967:Receptors, Gonadotropin / Q000302:isolation & purification* / Q000378:metabolism; D011974:Receptors, LH / Q000302:isolation & purification* / Q000378:metabolism
D006063:Chorionic Gonadotropin; D046911:Macromolecular Substances; D011967:Receptors, Gonadotropin; D011974:Receptors, LH; D000262:Adenylyl Cyclases