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. 1986 Oct 1;239(1):83–87. doi: 10.1042/bj2390083

Rat ovarian lutropin receptor is a transmembrane protein. Evidence for an Mr-20,000 cytoplasmic domain.

K P Keinänen, H J Rajaniemi
PMCID: PMC1147242  PMID: 3800987

Abstract

Membrane topography of the rat ovarian lutropin receptor was studied by two different approaches. Ovarian membrane preparation, labelled with 125I-labelled human choriogonadotropin in vivo, was subjected to extensive chymotryptic digestion. The soluble and membrane-bound radioactive complexes were cross-linked with glutaraldehyde, and analysed by SDS/polyacrylamide-gel electrophoresis and autoradiography. Chymotrypsin solubilized 70-75% of the radioactivity as Mr-96,000, Mr-74,000 and Mr-61,000 complexes, and decreased the size of the membrane-bound 125I-labelled human choriogonadotropin-receptor complex from Mr 130,000 to Mr 110,000. The Mr-110,000 complex was not observed when 0.1% Triton X-100 was present in the proteolytic digestion. Enrichment of inside-out-oriented plasma-membrane vesicles by concanavalin A affinity chromatography increased by 70% the fraction of radioactivity that remained in the membrane fraction after chymotrypsin treatment. Chymotrypsin also diminished the size of the membrane-bound unoccupied receptor from Mr 90,000 to Mr 70,000, as detected by ligand (125I-labelled human choriogonadotropin) blotting. These results suggest that the lutropin receptor is a transmembrane protein with a cytoplasmic domain of Mr 20,000 that is sensitive to proteolytic digestion in the inside-out-oriented plasma-membrane vesicles.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Dattatreyamurty B., Rathnam P., Saxena B. B. Isolation of the luteinizing hormone-chorionic gonadotropin receptor in high yield from bovine corpora lutea. Molecular assembly and oligomeric nature. J Biol Chem. 1983 Mar 10;258(5):3140–3158. [PubMed] [Google Scholar]
  2. Dufau M. L., Baukal A. J., Catt K. J. Hormone-induced guanyl nucleotide binding and activation of adenylate cyclase in the Leydig cell. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5837–5841. doi: 10.1073/pnas.77.10.5837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dufau M. L., Ryan D. W., Baukal A. J., Catt K. J. Gonadotropin receptors. Solubilization and purification by affinity chromatography. J Biol Chem. 1975 Jun 25;250(12):4822–4824. [PubMed] [Google Scholar]
  4. Gilman A. G. G proteins and dual control of adenylate cyclase. Cell. 1984 Mar;36(3):577–579. doi: 10.1016/0092-8674(84)90336-2. [DOI] [PubMed] [Google Scholar]
  5. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  6. Hwang J., Menon K. M. Characterization of the subunit structure of gonadotropin receptor in luteinized rat ovary. J Biol Chem. 1984 Feb 10;259(3):1978–1985. [PubMed] [Google Scholar]
  7. Ji I., Bock J. H., Ji T. H. Composition and peptide maps of cross-linked human choriogonadotropin-receptor complexes on porcine granulosa cells. J Biol Chem. 1985 Oct 15;260(23):12815–12821. [PubMed] [Google Scholar]
  8. Ji I., Yoo B. Y., Kaltenbach C., Ji T. H. Structure of the lutropin receptor on granulosa cells. Photoaffinity labeling with the alpha subunit in human choriogonadotropin. J Biol Chem. 1981 Nov 10;256(21):10853–10858. [PubMed] [Google Scholar]
  9. Kellokumpu S., Rajaniemi H. Involvement of plasma membrane enzymes in the proteolytic cleavage of luteinizing hormone receptor. Endocrinology. 1985 Feb;116(2):707–714. doi: 10.1210/endo-116-2-707. [DOI] [PubMed] [Google Scholar]
  10. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  11. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  12. Markkanen S. O., Rajaniemi H. J. Role of internalization and degradation in the removal of receptor-bound human chorionic gonadotropin in rat luteal cells in vivo. Endocrinology. 1980 Oct;107(4):1153–1161. doi: 10.1210/endo-107-4-1153. [DOI] [PubMed] [Google Scholar]
  13. Markkanen S., Töllikkö K., Jäskeläinen K., Rajaniemi H. Effect of radioiodination on the structural integrity, receptor and antibody-binding activity and circulatory behavior of human choriogonadotropin. Horm Res. 1980;12(1):32–45. doi: 10.1159/000179103. [DOI] [PubMed] [Google Scholar]
  14. Metsikkö M. K., Rajaniemi H. J. The hormone-binding unit of luteinizing hormone receptor. Biochem J. 1982 Nov 15;208(2):309–316. doi: 10.1042/bj2080309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Rajan V. P., Menon K. M. Evidence that dissociation, not intracellular degradation, is the major pathway for removal of receptor-bound 125I-human chorionic gonadotropin in cultured rat luteal cells. Biol Reprod. 1985 Aug;33(1):60–66. doi: 10.1095/biolreprod33.1.60. [DOI] [PubMed] [Google Scholar]
  16. Rapoport B., Hazum E., Zor U. Photoaffinity labeling of human chorionic gonadotropin-binding sites in rat ovarian plasma membranes. J Biol Chem. 1984 Apr 10;259(7):4267–4271. [PubMed] [Google Scholar]
  17. Rebois R. V., Omedeo-Sale F., Brady R. O., Fishman P. H. Covalent crosslinking of human chorionic gonadotropin to its receptor in rat testes. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2086–2089. doi: 10.1073/pnas.78.4.2086. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Walsh F. S., Barber B. H., Crumpton M. J. Preparation of inside-out vesicles of pig lymphocyte plasma membrane. Biochemistry. 1976 Aug 10;15(16):3557–3563. doi: 10.1021/bi00661a025. [DOI] [PubMed] [Google Scholar]
  19. Wimalasena J., Moore P., Wiebe J. P., Abel J., Jr, Chen T. T. The porcine LH/hCG receptor. Characterization and purification. J Biol Chem. 1985 Sep 5;260(19):10689–10697. [PubMed] [Google Scholar]

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