Identification of luteinizing hormone receptor binding inhibitor in bovine corpora lutea.
A 7000 g supernatant, obtained during the purification of luteinizing hormone (LH) receptor from bovine corpora lutea homogenate, was concentrated by ultrafiltration. The filtrate, containing < 50,000 molecular weight material, exhibited LH receptor binding inhibitor (LH-RBI) activity. The filtrate was ultrafiltered sequentially through Amicon PM-10, PM-30 and UM-2 filters to yield a LH-RBI-containing fraction in the higher molecular weight range of 30,000-10,000 and a LH-RBI-containing fraction in the lower molecular weight range of 10,000-1000. The higher molecular weight LH-RBI fraction was purified on Sephadex G-25 and the lower molecular weight LH-RBI fraction was purified on Sephadex G-50. Both the high- and the low-molecular-weight LH-RBI species inhibited the binding of 125I-labeled human chorionic gonadotropin (hCG) to bovine corpora lutea and to rat Leydig cell membrane receptors. Similarly, the production of testosterone by hCG-stimulated rat Leydig cells was inhibited in a dose-response manner by both the high- and the low-molecular-weight LH-RBI species. The LH-RBI activity in the low-molecular-weight species was stable at 4 degrees C for up to 6 months and at temperatures up to 90 degrees C for 15 mins, whereas the LH-RBI activity of the high-molecular-weight species was stable at 4 degrees C for 15 months and unstable at 60 degrees C after 15 min. The 7000 g supernatant provided a much-needed source to obtain larger than previously reported quantities of LH-RBI for isolation as well as for structure and function studies.
Department of Obstetrics and Gynecology, Cornell University Medical College, New York 10021.
P
P
Rathnam
S Q
SQ
Lin
B B
BB
Saxena
D000818:Animals; D002417:Cattle; D006063:Chorionic Gonadotropin / Q000378:metabolism; D003338:Corpus Luteum / Q000378:metabolism*; D005260:Female; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D007985:Leydig Cells / Q000378:metabolism; D008297:Male; D008970:Molecular Weight; D010455:Peptides / Q000032:analysis* / Q000302:isolation & purification; D011485:Protein Binding; D011869:Radioligand Assay; D051381:Rats; D011974:Receptors, LH / Q000037:antagonists & inhibitors* / Q000302:isolation & purification
D006063:Chorionic Gonadotropin; D036341:Intercellular Signaling Peptides and Proteins; C011898:LH receptor binding inhibitor; D010455:Peptides; D011974:Receptors, LH