足月时人胎盘细胞质20α-羟基类固醇脱氢酶（EC 1.1.1.149）的部分特性分析

Partial characterization of the cytoplasmic 20 alpha-hydroxysteroid dehydrogenase (EC 1.1.1.149) of the human placenta at term.

作者信息

Rabe T, Kiesel L, Runnebaum B

出版信息

J Steroid Biochem. 1982 Jun;16(6):737-43. doi: 10.1016/0022-4731(82)90029-2.

Abstract

The 20 alpha-hydroxysteroid dehydrogenase (20 alpha-HSDH) is a key enzyme in human fetal and maternal progesterone metabolism. In this paper, the cytoplasmic 20 alpha-HSDH of human term placenta is partially characterized in vitro. A 14-fold concentration of the 20 alpha-HSDH was prepared by ultracentrifugation and ammonium sulfate precipitation. The apparent Km values for the substrates progesterone (Km: 4.8 x 10(-5) M) and 20 alpha-DHP (Km: 6.2 x 10(-5) M) and for the cofactors NADPH (Km: 1.9 x 10(-4)) and NADH (Km: 2.6 x 10(-4)) were determined. The temperature optimum for the oxidation of 20 alpha-DHP is 40--50 degrees C. The pH optimum for the reduction of progesterone was found to be pH 6.2 and for the oxidation of 20 alpha-DHP pH 6.5. The addition of glycerol (3 M) to the incubation medium inhibited the conversion rate of 20 alpha-HSDH by 70%. No influence of EDTA could be found. Various bivalent metal ions (1--100 mM) showed a dose-dependent inhibition of 20 alpha-HSDH; a complete inhibition was achieved at 100 mM: Cu2+, Zn2+, Cd2+, Fe2+ and Ni2+.

摘要

20α-羟类固醇脱氢酶（20α-HSDH）是人类胎儿和母体孕酮代谢中的关键酶。本文在体外对足月人胎盘的细胞质20α-HSDH进行了部分特性研究。通过超速离心和硫酸铵沉淀制备了浓度提高14倍的20α-HSDH。测定了底物孕酮（Km：4.8×10⁻⁵ M）、20α-二氢孕酮（Km：6.2×10⁻⁵ M）以及辅因子NADPH（Km：1.9×10⁻⁴）和NADH（Km：2.6×10⁻⁴）的表观Km值。20α-二氢孕酮氧化的最适温度为40 - 50℃。发现孕酮还原的最适pH为6.2，20α-二氢孕酮氧化的最适pH为6.5。向孵育培养基中添加甘油（3 M）可使20α-HSDH的转化率降低70%。未发现EDTA有影响。各种二价金属离子（1 - 100 mM）对20α-HSDH表现出剂量依赖性抑制；在100 mM时可实现完全抑制：Cu²⁺、Zn²⁺、Cd²⁺、Fe²⁺和Ni²⁺。