Use of monoclonal antibodies to subunits of human chorionic gonadotropin to examine the orientation of the hormone in its complex with receptor.

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Abstract
Monoclonal antibodies were prepared aginst the .alpha.- and .beta.-subunits of human chorionic gonadotropin (hCG). Although all were selected on the basis of their ability to bind the intact hormone, each also bound 1 of the 2 subunits but not both. Using a solid phase double antibody system to measure the relative binding to sites on the surface of hCG, 4 of the 5 antibodies bound to different sites on the molecule. This information was correlated with the ability of each antibody to inhibit the biological activity of hCG. Of the 5 antibodies tested for the ability to inhibit hCG-induced stimulation of rat testes steroidogenesis in vitro, 2 proved to be potent inhibitors, whereas the other 3 had almost no effect. This inhibition of steroidogenesis was highly correlated with the ability of the antibodies to inhibit binding to testes homogenates. A scheme was derived that describes the relative binding positions of individual monoclonal antibodies and receptor on hCG. The purified monoclonal antibodies were iodinated and employed to evaluate which antigenic sites on hCG remained free in hCG-receptor complexes. Portions of the .beta.-subunit in hCG-receptor complexes were buried (i.e., failed to bind radiolabeled antibody), whereas other portions remained exposed (i.e., they bound radiolabeled antibody). Those antibodies that interacted with portions of hCG that became inaccessible in the receptor complex also blocked the biological actions of hCG, whereas those that interacted with exposed sites had little or no effect on activity. Although antibodies were not found to the .alpha.-subunit that would bind to the hormone-receptor complex, 1 of the 2 antibodies specific to .alpha.-subunit epitopes blocked the actions of the hormone. Both antigenic determinants on the .alpha.-subunit appeared to be lost after the hCG-receptor complex had formed. Each hCG subunit apparently participates in the hormone-receptor complex and portions of the .beta.-subunit project from the surface of the receptor.