G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Moscow Region, Russia.
J Steroid Biochem Mol Biol. 2012 Mar;129(1-2):47-53. doi: 10.1016/j.jsbmb.2011.09.008. Epub 2011 Oct 10.
Fast-growing strain of Mycobacterium sp. VKM Ac-1815D is capable of effective oxidizing of sterols (phytosterol, cholesterol, ergosterol) to androstenedione and other valuable 3-oxo-steroids. To elucidate the role of cholesterol oxidase in sterol catabolism by the strain, the choD gene has been cloned and sequenced. The deduced gene product (M(r) 63.5kDa) showed homologies over its entire length to a large number of proteins belonging to the InterPro-family EPR006076, which includes various FAD dependent oxidoreductases. The expression of choD in Escherichia coli was shown to result in the synthesis of membrane associated cholesterol oxidase. In addition to cholesterol, the enzyme oxidized β-sitosterol, dehydroepiandrosterone, ergosterol, pregnenolone, and lithocholic acid. Knock-out of choD in Mycobacterium sp. VKM Ac-1815D strain was obtained by the gene replacement technique. The mutant strain transformed sitosterol forming exclusively 3-keto-4-ene steroids with androstenedione as a major product, thus evidencing that choD knock out did not abrogate sterol A-ring oxidation. The results indicated that ChoD is not a critical enzyme responsible for modification of 3β-hydroxy-5-ene- to 3-keto-4-ene steroids in Mycobacterium sp. VKM Ac-1815D. Article from a special issue on steroids and microorganisms.
快速生长的分枝杆菌菌株 VKM Ac-1815D 能够有效地将甾醇（植物甾醇、胆固醇、麦角甾醇）氧化为雄烯二酮和其他有价值的 3-氧代甾族化合物。为了阐明胆固醇氧化酶在该菌株甾醇分解代谢中的作用，已经克隆和测序了 choD 基因。推断的基因产物（M(r)63.5kDa）在全长范围内与属于 InterPro 家族 EPR006076 的大量蛋白质具有同源性，其中包括各种依赖 FAD 的氧化还原酶。证明在大肠杆菌中表达 choD 会导致膜结合胆固醇氧化酶的合成。除了胆固醇，该酶还氧化 β-谷甾醇、脱氢表雄酮、麦角甾醇、孕烯醇酮和石胆酸。通过基因替换技术获得了分枝杆菌 VKM Ac-1815D 菌株中 choD 的缺失突变株。该突变株转化 sitosterol 仅形成雄烯二酮作为主要产物的 3-酮-4-烯甾体，从而证明 choD 敲除并未消除甾醇 A 环氧化。结果表明，ChoD 不是负责分枝杆菌 VKM Ac-1815D 中 3β-羟基-5-烯-至 3-酮-4-烯甾体修饰的关键酶。该文章来自于关于类固醇和微生物的特刊。
