ISOLATION OF 9 DIFFERENT BIOLOGICALLY AND IMMUNOLOGICALLY ACTIVE MOLECULAR VARIANTS OF BOVINE FOLLICULAR INHIBIN

被引:51
作者
GOOD, TEM
WEBER, PSD
IRELAND, JLH
PULASKI, J
PADMANABHAN, V
SCHNEYER, AL
LAMBERTMESSERLIAN, G
GHOSH, BR
MILLER, WL
GROOME, N
IRELAND, JJ
机构
[1] MICHIGAN STATE UNIV, DEPT ANIM SCI, MOLEC REPROD ENDOCRINOL LAB, E LANSING, MI 48824 USA
[2] UNIV MICHIGAN, DEPT PEDIAT, ANN ARBOR, MI 48109 USA
[3] UNIV MICHIGAN, REPROD SCI PROGRAM, ANN ARBOR, MI 48109 USA
[4] MASSACHUSETTS GEN HOSP, DEPT MED, REPROD ENDOCRINE UNIT, BOSTON, MA 02114 USA
[5] N CAROLINA STATE UNIV, DEPT BIOCHEM, RALEIGH, NC 27695 USA
[6] OXFORD BROOKES UNIV, MULTIPLE SCLEROSIS SOC PEPTIDE LAB, OXFORD, ENGLAND
关键词
D O I
10.1095/biolreprod53.6.1478
中图分类号
Q [生物科学];
学科分类号
07 ; 0710 ; 09 ;
摘要
A combination of immunoaffinity chromatography, SDS-PAGE, and electroelution was used to simultaneously isolate 0.36-4.65 mg of nine different molecular forms of inhibin (pro alpha(c)-29 kDa; fully processed 34 kDa; and large inhibin forms 49, 53, 58, 77, 88, 110, and > 160 kDa) from 0.675 L of bovine follicular fluid (bFF). Each inhibin form, except pro alpha(c)(1-26), cross-reacted with inhibin alpha(c)(1-26)- and beta(A)(82-114)-subunit-directed antibodies during immunoblot analysis. Pro alpha(c) cross-reacted only with alpha-subunit antibodies. The inhibin forms consisted of 22-, 29-, 49-, or 58-kDa alpha subunits and 17- or 58-kDa beta subunits. During cultures of ovine pituitary cells, a 5-ng/ml dose of each inhibin form (except pro alpha(c)) suppressed basal accumulation of FSH 30% to 50% but increased GnRH-induced LH release 40% to 248%. The various inhibin forms cross-reacted in a parallel fashion with standard curves generated during homologous and heterologous RIAs but with markedly different relative immunopotencies. In the RIAs, pro alpha(c) cross-reacted 3- to 18-fold more than the fully processed inhibin form. The fully processed and the seven different large forms of inhibin cross-reacted with different relative immunopotencies in a two-site dimer-specific ELISA. We concluded that 1) a combination of immunoaffinity extraction, SDS-PAGE, and electroelution simultaneously isolated relatively large amounts of highly enriched preparations of nine different molecular forms of immunologically and biologically active inhibin from bFF; 2) eight different dimeric forms of bovine inhibin may regulate both basal FSH and GnRH-induced LH secretion by the pituitary gland, and 3) eight or nine different molecular forms of inhibin cross-react with different relative immunopotencies in the two-site dimer-specific assay or RIAs.
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页码:1478 / 1488
页数:11
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