Akao T, Akao T, Kobashi K
Faculty of Pharmaceutical Sciences, Toyama Medical and Pharmaceutical University, Japan.
Biochim Biophys Acta. 1990 Feb 6;1042(2):241-6. doi: 10.1016/0005-2760(90)90015-p.
Glycyrrhetic acid, derived from a main component of liquorice, was converted to 3-ketoglycyrrhetic acid reversibly by rat liver homogenates in the presence of NADPH or NADP+. Glycyrrhetic acid-oxidizing and 3-ketoglycyrrhetic acid-reducing activities were localized in microsomes among the subcellular fractions of rat liver. Glycyrrhetic acid-oxidizing activity and 3-ketoglycyrrhetic acid-reducing activities showed pH optima at 6.3 and 8.5, respectively, and required NADP+ or NAD+ and NADPH or NADH, respectively, indicating that these activities were due to glycyrrhetinate dehydrogenase. The dehydrogenase was not solubilized from the membranes by the treatment with 1 M NaCl or sonication, indicating that the enzyme is a membrane component. The dehydrogenase was solubilized with detergents such as Emalgen 913, Triton X-100 and sodium cholate, and then separated from 3 beta-hydroxysteroid dehydrogenase (5 beta-androstan-3 beta-ol-17-one-oxidizing activity) by butyl-Toyopearl 650 M column chromatography. Partially purified enzyme catalyzed the reversible reaction between glycyrrhetic acid and 3-ketoglycyrrhetic acid, but was inactive toward 3-epiglycyrrhetic acid and other steroids having the 3 beta-hydroxyl group. The enzyme required NADP+ and NADPH for the highest activities of oxidation and reduction, respectively, and NAD+ and NADH for considerable activities, similar to the results with microsomes. From these results the enzyme is defined as glycyrrhetinate dehydrogenase, being quite different from 3 beta-hydroxysteroid dehydrogenase of Ruminococcus sp. from human intestine, which is active for both glycyrrhetic acid and steroids having the 3 beta-hydroxyl group.
甘草次酸是从甘草的主要成分中提取得到的，在存在烟酰胺腺嘌呤二核苷酸磷酸（NADPH）或烟酰胺腺嘌呤二核苷酸（NADP+）的情况下，大鼠肝脏匀浆可使其可逆地转化为3-酮基甘草次酸。在大鼠肝脏的亚细胞组分中，甘草次酸氧化活性和3-酮基甘草次酸还原活性定位于微粒体中。甘草次酸氧化活性和3-酮基甘草次酸还原活性的最适pH分别为6.3和8.5，分别需要NADP+或NAD+以及NADPH或NADH，这表明这些活性是由甘草次酸脱氢酶引起的。用1M氯化钠处理或超声处理并不能使该脱氢酶从膜上溶解下来，这表明该酶是一种膜成分。该脱氢酶可用诸如乳化剂913、聚山梨醇酯80和胆酸钠等去污剂溶解，然后通过丁基琼脂糖650M柱色谱法与3β-羟基类固醇脱氢酶（5β-雄甾烷-3β-醇-17-酮氧化活性）分离。部分纯化的酶催化甘草次酸和3-酮基甘草次酸之间的可逆反应，但对3-表甘草次酸和其他具有3β-羟基的类固醇无活性。该酶分别需要NADP+和NADPH以达到最高的氧化和还原活性，需要NAD+和NADH以达到相当的活性，这与微粒体的结果相似。根据这些结果，该酶被定义为甘草次酸脱氢酶，与人肠道中瘤胃球菌属的3β-羟基类固醇脱氢酶有很大不同，后者对甘草次酸和具有3β-羟基的类固醇均有活性。
