Identification and characterization of a 20β-HSDH from the anaerobic gut bacterium Butyricicoccus desmolans ATCC 43058.
Identification and characterization of a 20β-HSDH from the anaerobic gut bacterium Butyricicoccus desmolans ATCC 43058.
复制标题
DOI：
10.1194/jlr.m074914
复制
发表时间：
2017-05
影响因子：
6.5
通讯作者：
Ridlon JM
中科院分区：
文献类型：
--
作者：
Devendran S;Méndez-García C;Ridlon JM
已知胃肠道微生物群的成员将糖皮质激素转化为雄甾烷，雄甾烷随后被肠道微生物群或宿主组织的其他成员转化为有效的雄激素。先前已经报道了Butyricicoccus desmolans和Clostridium cadaveris的类固醇-17，20-碳链酶和20β-羟基类固醇脱氢酶（HSDH）活性，其负责从皮质醇形成雄甾烷;然而，编码这些酶的基因尚未报道。在这项工作中，我们确定和定位的基因编码20β-HSDH在两个B。desmolans和C.尸体B的20β-HSDH。desmolans异源过表达和纯化大肠杆菌。该酶为同源四聚体，亚基分子量为33.8 ± 3.7kDa。r20β-HSDH以（20β-dihydro）cortisol和NAD（H）为底物，其还原方向的最适pH为9.0，氧化方向的最适pH为7.0-7.5。皮质醇是首选底物，Km为0.80 ± 0.06 μM; Vmax为30.36 ± 1.97 μmol·min-1; Kcat为607 ± 39 μmol·μM-1·min-1; Kcat/Km为760 ± 7.67。B 20β-HSDH基因的系统发育分析desmolans表明20β-HSDH存在于几种双歧杆菌属中，其中1株表达20β-HSDH活性。值得注意的是，我们还确定了一种新的类固醇-17，20-碳链酶-阐述细菌，Propionimicrobium lymphophilum，一个正常的居民的泌尿道。
Members of the gastrointestinal microbiota are known to convert glucocorticoids to androstanes, which are subsequently converted to potent androgens by other members of the gut microbiota or host tissues. Butyricicoccus desmolans and Clostridium cadaveris have previously been reported for steroid-17,20-desmolase and 20β-hydroxysteroid dehydrogenase (HSDH) activities that are responsible for androstane formation from cortisol; however, the genes encoding these enzymes have yet to be reported. In this work, we identified and located a gene encoding 20β-HSDH in both B. desmolans and C. cadaveris. The 20β-HSDH of B. desmolans was heterologously overexpressed and purified from Escherichia coli. The enzyme was determined to be a homotetramer with subunit molecular mass of 33.8 ± 3.7 kDa. The r20β-HSDH displayed pH optimum in the reductive direction at pH 9.0 and in the oxidative direction at pH 7.0–7.5 with (20β-dihydro)cortisol and NAD(H) as substrates. Cortisol is the preferred substrate with Km, 0.80 ± 0.06 μM; Vmax, 30.36 ± 1.97 μmol·min−1; Kcat, 607 ± 39 μmol·μM−1·min−1; Kcat/Km, 760 ± 7.67. Phylogenetic analysis of the 20β-HSDH from B. desmolans suggested that the 20β-HSDH is found in several Bifidobacterium spp., one of which was shown to express 20β-HSDH activity. Notably, we also identified a novel steroid-17,20-desmolase-elaborating bacterium, Propionimicrobium lymphophilum, a normal inhabitant of the urinary tract.