Center of Innovations and Technologies "Biological Active Compounds and Their Applications", Russian Academy of Sciences, Moscow 119991, Russian Federation; G.K.Skryabin Institute of Biochemistry & Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Moscow Region, Russian Federation.
J Steroid Biochem Mol Biol. 2013 Nov;138:41-53. doi: 10.1016/j.jsbmb.2013.02.016. Epub 2013 Mar 6.
A comparative genome analysis of Mycobacterium spp. VKM Ac-1815D, 1816D and 1817D strains used for efficient production of key steroid intermediates (androst-4-ene-3,17-dione, AD, androsta-1,4-diene-3,17-dione, ADD, 9α-hydroxy androst-4-ene-3,17-dione, 9-OH-AD) from phytosterol has been carried out by deep sequencing. The assembled contig sequences were analyzed for the presence putative genes of steroid catabolism pathways. Since 3-ketosteroid-9α-hydroxylases (KSH) and 3-ketosteroid-Δ(1)-dehydrogenase (Δ(1) KSTD) play key role in steroid core oxidation, special attention was paid to the genes encoding these enzymes. At least three genes of Δ(1) KSTD (kstD), five genes of KSH subunit A (kshA), and one gene of KSH subunit B of 3-ketosteroid-9α-hydroxylases (kshB) have been found in Mycobacterium sp. VKM Ac-1817D. Strains of Mycobacterium spp. VKM Ac-1815D and 1816D were found to possess at least one kstD, one kshB and two kshA genes. The assembled genome sequence of Mycobacterium sp. VKM Ac-1817D differs from those of 1815D and 1816D strains, whereas these last two are nearly identical, differing by 13 single nucleotide substitutions (SNPs). One of these SNPs is located in the coding region of a kstD gene and corresponds to an amino acid substitution Lys (135) in 1816D for Ser (135) in 1815D. The findings may be useful for targeted genetic engineering of the biocatalysts for biotechnological application.
对用于从植物甾醇高效生产关键甾体中间体（雄甾-4-烯-3,17-二酮、AD、雄甾-1,4-二烯-3,17-二酮、ADD、9α-羟基雄甾-4-烯-3,17-二酮、9-OH-AD）的 Mycobacterium spp. VKM Ac-1815D、1816D 和 1817D 菌株进行了比较基因组分析，该分析是通过深度测序完成的。对组装的连续序列进行了分析，以确定是否存在甾体分解代谢途径的推定基因。由于 3-酮甾体-9α-羟化酶（KSH）和 3-酮甾体-Δ(1)-脱氢酶（Δ(1) KSTD）在甾体核心氧化中起关键作用，因此特别关注编码这些酶的基因。在 Mycobacterium sp. VKM Ac-1817D 中发现了至少三个 Δ(1) KSTD（kstD）基因、五个 KSH 亚单位 A（kshA）基因和一个 3-酮甾体-9α-羟化酶 KSH 亚单位 B（kshB）基因。Mycobacterium spp. VKM Ac-1815D 和 1816D 菌株被发现至少具有一个 kstD、一个 kshB 和两个 kshA 基因。Mycobacterium sp. VKM Ac-1817D 的组装基因组序列与 1815D 和 1816D 菌株的序列不同，而后两者几乎相同，仅相差 13 个单核苷酸取代（SNP）。这些 SNP 中的一个位于 kstD 基因的编码区，对应于 1816D 中的 Ser（135）替换为 1815D 中的 Lys（135）。这些发现可能有助于针对生物技术应用的生物催化剂进行靶向遗传工程。
