Chicz R M, Nakamura R M, Goebelsmann U, Campeau J D, Tonetta S A, Frederick J J, diZerega G S
J Steroid Biochem. 1985 Nov;23(5A):663-8. doi: 10.1016/0022-4731(85)90019-6.
A heat- and trypsin-labile follicular fluid protein (FRP) extracted from both human and porcine follicular fluid has been shown to modulate ovarian steroidogenesis. To further investigate the effects of FRP, its effect on the kinetics of 3 beta-hydroxysteroid dehydrogenase activity (3 beta-HSD) was evaluated in cell-free microsomal preparations from human placenta. Test fractions of follicular fluid protein were preincubated with placental microsomes followed by the addition of various substrate concentrations (pregnenolone + NAD). Subsequent progesterone formation was interpreted as the velocity of the reaction. The 50% inhibitory dose (ID50) of FRP for 3 beta-HSD for the three substrate concentrations was 300 micrograms/ml. Although a clear decrease in 3 beta-HSD activity typically occurred after pre-incubation with 730 micrograms/ml of FRP, a paradoxical augmentation in 3 beta-HSD activity was present with the lower concentrations of FRP (10-30 micrograms/ml) and the more concentrated microsomal preparations. Double reciprocal plots of these reactions demonstrated a Km for 3 beta-HSD of 1.8-2.1 X 10(-6) M. Analysis of all reactions was found to be consistent with a noncompetitive mode of enzyme inhibition with an apparent Ki of 120 ng/ml or approximately 10(-8) M assuming a mol. wt of 16,000 Daltons for FRP. This derived Ki for FRP is within the biological concentration of FRP in follicular fluid.
从人及猪卵泡液中提取的一种对热和胰蛋白酶敏感的卵泡液蛋白（FRP）已被证明可调节卵巢类固醇生成。为进一步研究FRP的作用，在人胎盘无细胞微粒体制剂中评估了其对3β-羟基类固醇脱氢酶活性（3β-HSD）动力学的影响。将卵泡液蛋白的测试组分与胎盘微粒体预孵育，随后加入不同浓度的底物（孕烯醇酮+NAD）。随后的孕酮生成被视为反应速度。对于三种底物浓度，FRP对3β-HSD的50%抑制剂量（ID50）为300微克/毫升。尽管在与730微克/毫升的FRP预孵育后，3β-HSD活性通常会明显降低，但在较低浓度的FRP（10 - 30微克/毫升）和更浓缩的微粒体制剂中，3β-HSD活性却出现了反常的增强。这些反应的双倒数图显示3β-HSD的Km为1.8 - 2.1×10⁻⁶ M。所有反应分析均表明其符合非竞争性酶抑制模式，假设FRP的分子量为16000道尔顿，其表观Ki为120纳克/毫升或约10⁻⁸ M。FRP的这种推导Ki值在卵泡液中FRP的生物浓度范围内。
