title: Comparative analysis of genes encoding key steroid core oxidation enzymes in fast-growing Mycobacterium spp. strains.
abstract: 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.
authors.affiliation: 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. Electronic address: bragory@yandex.ru.
authors.forename: E Yu
authors.initials: EY
authors.lastname: Bragin
authors.forename: V Yu
authors.initials: VY
authors.lastname: Shtratnikova
authors.forename: D V
authors.initials: DV
authors.lastname: Dovbnya
authors.forename: M I
authors.initials: MI
authors.lastname: Schelkunov
authors.forename: Yu A
authors.initials: YA
authors.lastname: Pekov
authors.forename: S G
authors.initials: SG
authors.lastname: Malakho
authors.forename: O V
authors.initials: OV
authors.lastname: Egorova
authors.forename: T V
authors.initials: TV
authors.lastname: Ivashina
authors.forename: S L
authors.initials: SL
authors.lastname: Sokolov
authors.forename: V V
authors.initials: VV
authors.lastname: Ashapkin
authors.forename: M V
authors.initials: MV
authors.lastname: Donova
mesh_terms: D000730:Androstadienes / Q000378:metabolism; D000735:Androstenedione / Q000031:analogs & derivatives / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism; D009161:Mycobacterium / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D010840:Phytosterols / Q000378:metabolism
chemical_list: D000730:Androstadienes; D001426:Bacterial Proteins; D010840:Phytosterols; C006573:1,4-androstadiene-3,17-dione; D000735:Androstenedione; C076717:9-hydroxy-4-androstene-3,17-dione; D006899:Mixed Function Oxygenases; D010088:Oxidoreductases
keywords: 2,3-dehydroxyphenyl dioxygenase; 2-enoyl acyl-CoA hydratase; 2-hydroxypenta-2,4-dienoate hydratase; 3,4-dihydroxy-9,10-secoandrosta-1,3,5(10)-triene-9,17-dione 4,5-dioxygenase; 3-hydroxy-9,10-secoandrosta-1,3,5(10)-triene-9,17-dione monooxygenase; 3-hydroxy-9,10-secoandrosta-1,3,5(10)-triene-9,17-dione monooxygenase subunit; 3-ketosteroid-9α-hydroxylase; 3-ketosteroid-Δ(1)-dehydrogenase; 3β-hydroxysteroid-dehydrogenase; 4,5:9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-diene-4-oate hydrolase; 4-hydroxy-2-oxovalerate aldolase; 9-OH-AD; 9α-hydroxy androst-4-ene-3,17-dione; AD; ADD; Androst-1,4-diene-3,17-dione; Androst-4-ene-3,17-dione; BWA; Broadband-Wheeler Aligner; CTAB; ChoX; ChoX(D,E); EchA19; FAD; FadA5; FadD17; FadD19; FadE26; FadE27; FadE28; Genome sequencing; HSD; HTH-type transcriptional repressor; HsaA; HsaAB; HsaB; HsaC; HsaD; HsaE; HsaF; HsaG; Hsd4A; Hsd4B; KSH; KshA; KshB; KstR; KstR2; Ltp2; Ltp3; Ltp4; Mycobacterium; ORFs; PWM; Phytosterol; SNP; Steroid bioconversion; TesB; YrbE4A; YrbE4B; acetaldehyde dehydrogenase; acetyl-CoA acetyltransferase; acyl-CoA dehydrogenase; acyl-CoA synthetase; acyl-CoA thioesterase II; androst-4-ene-3,17-dione; androsta-1,4-diene-3,17-dione; base pair; bp; cetyl trimethyl ammonium bromide; cholesterol oxidase; enoyl-CoA hydratase; flavin adenine dinucleotide; hydroxysteroid dehydrogenase; integral membrane protein; lipid transfer protein 4 (keto acyl-CoA thiolase); lipid-transfer protein 2; lipid-transfer protein 3 (acetyl-CoA acetyltransferase); open reading frames; position weight matrix; single nucleotide substitution; subunit A of 3-ketosteroid-9α-hydroxylase; subunit B of 3-ketosteroid-9α-hydroxylases; Δ(1) KSTD