Molecular cloning and expression of the ovine testicular follicle stimulating hormone receptor

Yarney, T.A.; Sairam, M.R.; Khan, Haseena; Ravindranath, N. H.; Payne, S.; Seidah, Nabil G.

doi:10.1016/0303-7207(93)90127-6

Cited by 64 publications

(32 citation statements)

References 31 publications

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“…Partial structural analysis of the N-terminal region of the RP-HPLC material revealed the sequence MRGSHHHHHHTDPXH HRLXHXSNGVF. As the protein was not reduced and alkylated, X represents cysteine, which is consistent with the sequence predicted by cDNA cloning (Khan et al 1993). The 14th residue in the sequence represents the first amino acid of the mature FSH-R protein (Yarney et al 1993).…”

Section: Structure Of the Recombinant Proteinsupporting

confidence: 66%

“…The insert cDNA of 151A1 clone (Khan et al 1993), omitting the signal sequence (of first 17 amino acids) and any of the 3 -non-coding region, was generated by PCR using a forward primer carrying a BamH1 restriction site (underlined). 5 -ATAATTGGATCCCTGCCACCATCGA-3 .…”

Section: Construction Of the 151a1 Expression Plasmid And Purificatiomentioning

confidence: 99%

“…The effect of rabbit antibody to recombinant 151A1 protein was analyzed by pretreating HEK293 cells expressing the full-length FSH-R cDNA (Sairam et al 1996a, Yarney et al 1993. Cyclic AMP response to added FSH was quantitated by radioimmunoassay (Sairam et al 1996a).…”

Section: Effect Of Antibody To Recombinant Fsh-r Proteinmentioning

confidence: 99%

“…The FSH receptor (FSH-R) cloned from several species (Gromoll et al 1993, Kelton et al 1992, Minegish et al 1991, Sprengel et al 1990, Yarney et al 1993) is similar to LH and TSH receptors (Segaloff & Ascoli 1993), belonging in a subfamily of G-protein-coupled receptors that differ from others by having a large extracellular domain composed of more than 300 amino acid residues. This region of the receptor is believed to contribute binding sites to accommodate unique conformations presented by each of the dimeric hormones (Combarnous 1992, Sairam 1983, Segaloff & Ascoli 1993.…”

Section: Introductionmentioning

confidence: 99%

“…Several different alternately spliced forms of testicular FSH-R cDNA have been cloned from human (Gromoll et al 1992, Kelton et al 1992, ovine (Khan et al 1993, Sairam et al 1996a, bovine (M R Sairam, unpublished data) and mouse (O'Shaughnessy & Dudley 1993) sources. However, their functions and structural implications have remained a mystery.…”

Section: Introductionmentioning

confidence: 99%

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Recognition of follicle stimulating hormone (alpha-subunit) by a recombinant receptor protein domain coded by an alternately spliced mRNA and expressed in Escherichia coli

Khan¹,

Jiang²,

Jayashree³

et al. 1997

Journal of Molecular Endocrinology

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To assess the functional significance of putative proteins encoded by alternately spliced mRNA of the sheep testicular FSH receptor, a short form cDNA comprising of the first four exons (117 residues mature protein) was engineered for expression in Escherichia coli. The expressed protein of molecular mass 15 kDa was purified to homogeneity and verified by reaction with an antibody against a synthetic peptide sequence unique to the amino (N)-terminal region FSH receptor. The purified FSH receptor domain protein bound 125 I-labeled hFSH in a ligand blot on polyvinylidine difluoride membranes. Further analyses by slot blot revealed high affinity of the immobilized protein with significant reaction at 10 pmol. As the immobilized receptor protein also reacted with structurally related hormones ( 125 I-labeled LH/ 125 I-labeled human chorionic gonadotropin), we confirmed that interaction most probably occurred via the common -subunit of these glycoprotein hormones. Our results reveal that this N-terminal portion of the FSH receptor contain(s) major site(s) for hormone recognition that could be mediated via the -subunit. A rabbit antibody to the receptor inhibited FSH action in receptor bearing cells, revealing the utility of such recombinant FSH receptor protein(s) for modulation of hormone action.

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Section: Structure Of the Recombinant Proteinsupporting

confidence: 66%

Section: Construction Of the 151a1 Expression Plasmid And Purificatiomentioning

confidence: 99%