Subunit structure of the follitropin (FSH) receptor. Photoaffinity labeling of the membrane-bound receptor follitropin complex in situ

Journal Article · Fri Nov 15 04:00:00 EST 1985 · J. Biol. Chem.; (United States)

OSTI ID:6292096

Smith, R A; Branca, A A; Reichert, Jr, L E

Human follicle-stimulating hormone (hFSH) was acylated with N-hydroxysuccinimidyl-4-azidobenzoate (HSAB) and radioiodinated (55 microCi/micrograms) for use as a photoaffinity probe to investigate the subunit structure of the FSH receptor in calf testis. After incubation with the photoaffinity probe and photolysis with UV light, the cross-linked hormone-receptor complex was solubilized from the membrane and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the presence and absence of the reducing agent dithiothreitol. Autoradiography of the polyacrylamide gels revealed two major bands, 64 kDa and 84 kDa. These were equivalent in molecular mass to those observed in a previous study in which performed hormone-receptor complexes were solubilized with detergent prior to formation of covalent cross-linkages through the use of homobifunctional cross-linking reagents. Reduction with dithiothreitol resulted in the loss of radioactivity from the 84-kDa band with a concomitant increase in the intensity of the 64-kDa band. Since dithiothreitol increases the dissociation of intact radioiodinated azidobenzoyl-FSH into subunits, it is suggested that the conversion of the 84-kDa band to the 64-kDa band by dithiothreitol is due to the loss of non-cross-linked hFSH subunit from the 84-kDa band and that the two bands observed after photoaffinity labeling arise from covalent bond formation between hFSH and a receptor subunit having a relative molecular weight (Mr) of 48,000. In addition to the predominant photolabeling of the receptor to yield the 64-kDa and 84-kDa bands, several other, less intense bands (54 kDa, 76 kDa, 97 kDa, and 116 kDa) were also consistently observed on autoradiographs.

• Research Organization: Albany Medical College of Union Univ., NY
• OSTI ID: 6292096
• Journal Information: J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 26; ISSN JBCHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACYLATION
AFFINITY
ANIMALS
AUTORADIOGRAPHY
AZIDES
BODY
CATTLE
CELL CONSTITUENTS
CELL MEMBRANES
CENTRIFUGATION
CHEMICAL REACTIONS
COVALENCE
CROSS-LINKING
DECOMPOSITION
DOMESTIC ANIMALS
ELECTROPHORESIS
FSH
GONADOTROPINS
GONADS
HORMONES
IODINE ISOTOPES
ISOTOPES
LABELLING
MALE GENITALS
MAMMALS
MEMBRANES
MOLECULAR STRUCTURE
MOLECULAR WEIGHT
NITROGEN COMPOUNDS
ORGANS
PEPTIDE HORMONES
PHOTOCHEMICAL REACTIONS
PHOTOLYSIS
PITUITARY HORMONES
POLYMERIZATION
RECEPTORS
RUMINANTS
SEPARATION PROCESSES
TESTES
VERTEBRATES