Biochemical and physiologic characterization of follicle regulatory protein: a paracrine regulator of folliculogenesis.
Further purification of a porcine follicular fluid fraction, referred to as follicle regulatory protein, that inhibits granulosa cell aromatase was performed and the results of in vitro bioassays with these highly purified reagents are reported. The 0% to 35% saturated ammonium sulfate extract of porcine follicular fluid was percolated through an orange A dye matrex gel column and the bound fraction was eluted. Further purification of 0% to 35% orange A-bound fraction of porcine follicular fluid was performed by anion exchange chromatography with the use of the Mono Q column. Mono Q eluents containing follicle regulatory protein activity were injected onto a Mono P hydrogen ion-exchange column. Samples obtained from Mono P chromatography were injected onto preparative and analytical scale gel exclusion columns. Eluent fractions in the apparent molecular weight of 16,000 daltons were tested for aromatase inhibition. Throughout each step, parallelism of an aromatase inhibitor was apparent in both a cell-free microsomal assay and a granulosa cell assay. Follicle regulatory protein, purified about 6666-fold from the orange A-bound fraction of porcine follicular fluid, had a 50% inhibitory concentration of 25 ng/ml for granulosa cell aromatase activity.
D000818:Animals; D047072:Aromatase Inhibitors; D001391:Azo Compounds; D001681:Biological Assay / Q000379:methods; D002850:Chromatography, Gel / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D004396:Coloring Agents; D005260:Female; D005498:Follicular Phase; D006107:Granulosa Cells / Q000187:drug effects / Q000201:enzymology / Q000502:physiology; D066298:In Vitro Techniques; D036341:Intercellular Signaling Peptides and Proteins; D008861:Microsomes / Q000187:drug effects / Q000201:enzymology; D009281:Naphthalenes; D006080:Ovarian Follicle / Q000502:physiology*; D010455:Peptides / Q000302:isolation & purification / Q000494:pharmacology / Q000502:physiology*; D013552:Swine; D014227:Triazines
D047072:Aromatase Inhibitors; D001391:Azo Compounds; D004396:Coloring Agents; D036341:Intercellular Signaling Peptides and Proteins; D009281:Naphthalenes; D010455:Peptides; D014227:Triazines; C018355:intraovarian peptides; C048536:matrex gel orange A