Warneck H M, Seitz H U
Universität Tübingen, Botanisches Institut, Allgemeine Botanik und Pflanzenphysiologie, Bundesrepublik Deutschland.
Z Naturforsch C J Biosci. 1990 Sep-Oct;45(9-10):963-72. doi: 10.1515/znc-1990-9-1007.
A 3 beta-hydroxysteroid oxidoreductase was isolated and characterized in the microsomes of Digitalis lanata cell cultures. The enzyme catalyzes the conversion of 5 alpha-pregnane-3,20-dione to 5 alpha-pregnan-3 beta-ol-20-one and requires NAD(P)H2. The enzyme was found to have a pH optimum of 8.0. The reaction had an optimum incubation temperature of 25 degrees C with linear reduction for the first 4 h, reaching maximum enzyme activity after 7 h. Substrate kinetics for 5 alpha-pregnane-3,20-dione and NADPH2 resulted in apparent Km-values of 18.5-20 microM for 5 alpha-pregnane-3,20-dione and 50-120 microM for the co-substrate NADPH2. In order to localize 3 beta-hydroxysteroid oxidoreductase differential centrifugation as well as linear sucrose density gradient centrifugation were performed. The results obtained lead to the conclusion that 3 beta-hydroxysteroid oxidoreductase is not associated with a single cell compartment, but consists of a major soluble part and a markedly smaller part of endoplasmic reticulum-associated activity.
从毛花洋地黄细胞培养物的微粒体中分离并鉴定了一种3β-羟基类固醇氧化还原酶。该酶催化5α-孕烷-3,20-二酮转化为5α-孕烷-3β-醇-20-酮，反应需要NAD(P)H2。发现该酶的最适pH为8.0。反应的最适孵育温度为25℃，最初4小时呈线性还原，7小时后达到最大酶活性。5α-孕烷-3,20-二酮和NADPH2的底物动力学导致5α-孕烷-3,20-二酮的表观Km值为18.5 - 20μM，共底物NADPH2的表观Km值为50 - 120μM。为了定位3β-羟基类固醇氧化还原酶，进行了差速离心以及线性蔗糖密度梯度离心。所得结果得出结论，3β-羟基类固醇氧化还原酶不与单个细胞区室相关，而是由主要的可溶性部分和明显较小的内质网相关活性部分组成。
