BIOCHEMICAL STUDIES ON THE LH-HCG RECEPTOR

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A receptor preparation specific to luteinizing hormone and to human chorionic gonadotropin has been isolated from bovine corpora lutea and is available in our laboratory in sufficient quantities to permit biochemical, structural and immunological studies proposed here. Subunits of the receptor of 85,000 and 38,000 molecular weight are also available in the purified form. The isolated receptor and its subunits are largely protein in nature with minor moieties of carbohydrates and lipids. We propose to perform recombination experiments to confirm that the 85K and 38K fragments are disulfide-linked subunits of the receptor molecule. Reconstitution experiments, where receptors are taken out of the biological membrane and put back into an artificial membrane environment will also be performed for the LH-hCG receptor using phospholipids. We will characterize the protein moieties by SDS-gel electrophoresis, ultracentrifugal analyses, peptide mapping, end group analyses and partial amino acid sequence determination. A library of monoclonal antibodies to the LH-hCG receptor and its subunits will be produced. The antibodies will be purified and mapped for their specificities. The monoclonal antibodies will be utilized in structural analyses of the receptor; in the immunocytochemical localization of the receptor; to follow the biochemical fate of the hormone-receptor complex in the cell. The monoclonal antibodies will be immobolized on a solid phase and used to characterize receptor fractions by affinity chromatography. The monoclonal antibodies will also be used in the cloning of genetic material. The cloned DNA sequences will be utilized to alternatively confirm the sequence of the receptor and subunits. The monoclonal antibodies can be used for the development of more specific assays of the receptor. Monoclonal antibodies, because of their selectivity, specificity and unlimited productibility, can lend themselves for clinical use when coupled to a drug. These studies are vital to understand the mechanism of hormone action at the molecular level during normal and abnormal gonadal function.