A decapeptide corresponding to the partial amino acid sequence of a high molecular weight human FSH receptor-binding inhibitor is a specific inhibitor of FSH binding.
We previously reported purification of a protein (approximately equal to 57 kDa) from human follicular fluid having FSH binding inhibitory (FSH-BI) activity. Purified hFSH-BI was cleaved with cyanogen bromide and trypsin. The resulting peptide fragments were separated by HPLC and sequence information for individual fragments was obtained. A ten amino acid sequence of hFSH-BI derived from this procedure was identified, and a corresponding peptide amide (BI-10) was synthesized and utilized for further study. A protein database search revealed no significant identity between this decapeptide and other known proteins. We examined the ability of BI-10 to inhibit binding of 125I-hFSH to FSH-receptor enriched bovine testes membranes utilizing a radioligand receptor assay (RRA). BI-10 inhibited binding of 125I-hFSH to its receptor in a concentration-related manner, with an ED50 of 300 microM. BI-10 had no effect on 125I-hCG binding to receptor even at concentrations up to 1000 microM, suggesting that the effect of BI-10 was specific for the interaction between FSH and its receptor. To assess bioactivity of BI-10, we investigated its effect on FSH-stimulated conversion of androstenedione to estradiol by rat Sertoli cells in primary culture in vitro. Inhibition of FSH-stimulated estradiol synthesis (FSH antagonist activity) was significant at a BI-10 concentration of 1000 microM. BI-10 also significantly inhibited FSH-stimulated cAMP accumulation in primary cultures of Sertoli cells when examined at the same concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
Lee
D W
DW
Division of Endocrinology and Metabolism, Albany Medical College, NY 12208, USA.
Grasso
P
P
Leng
N
N
Izquierdo
R
R
Crabb
J W
JW
Deziel
M R
MR
Reichert
L E
LE
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D000242:Cyclic AMP / Q000032:analysis; D004958:Estradiol / Q000032:analysis; D005260:Female; D005640:Follicle Stimulating Hormone / Q000378:metabolism*; D015571:Follicular Fluid / Q000737:chemistry; D006020:Glycopeptides / Q000138:chemical synthesis / Q000494:pharmacology*; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011863:Radioimmunoassay; D011869:Radioligand Assay; D011962:Receptors, FSH / Q000737:chemistry / Q000378:metabolism*; D017421:Sequence Analysis
D002352:Carrier Proteins; C484018:FSH-BI protein, human; D006020:Glycopeptides; D010446:Peptide Fragments; D011962:Receptors, FSH; D004958:Estradiol; D005640:Follicle Stimulating Hormone; D000242:Cyclic AMP