Gonadotropin receptors with specificity, high affinity and low capacity for luteinizing hormone and human chorionic gonadotropin (hCG) have been identified in rat luteal cells. To investigate the nature of the receptor, we have employed disuccinimidyl suberate, a cross-linker noncleavable by reducing agents, and dithiobis(succinimidyl propionate), a cleavable cross-linker, to covalently cross-link the 125I-hCG·receptor complex. The molecular weight of 125I-hCG-linked receptor complex and the receptor subunit structure were determined by electrophoresis in either 10 or 4.5% acrylamide in the presence of 0.1% sodium dodecyl sulfate with or without reducing agents. Autoradiographic analysis of the 125I-hCG-linked receptor separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing condition revealed a single labeled band corresponding to M(r) = 305,000 ± 15,000. However, electrophoresis performed in the presence of 50 nM dithiothreitol and 2% β-mercaptoethanol resulted in the appearance of four labeled bands corresponding to M(r) = 105,000 ± 4,000, 96,000 ± 5,000, 74,000 ± 4,000, and 62,000 ± 4,000 concomitant with the loss of the labeled band in the M(r) = 305,000 region. Further experiments demonstrated that these four labeled bands were derived from the same molecular species. In addition, the 125I-hCG-linked receptor in the absence of reducing agent was not dissociated into subunits even by treatment with strong denaturing agent (8 M urea). The appearance of the cross-linked 125I-hCG·receptor was effectively inhibited by the unlabeled β-subunit of hCG, intact hCG, and luteinizing hormone and partially inhibited by the α-subunit of hCG but not by choleratoxin, gonadotropin-releasing hormone, insulin or bovine serum albumin. These data suggest that 1) the hCG/luteinizing hormone receptor is an oligomeric complex linked by disulfide bonds and 2) that under reducing conditions, the oligomeric receptor dissociates into four nonidentical subunits.
|Number of pages||8|
|Journal||Journal of Biological Chemistry|
|Publication status||Published - 1984|
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