title: Bioconversion of 4-androstene-3,17-dione to androst-1,4-diene-3,17-dione by recombinant Bacillus subtilis expressing ksdd gene encoding 3-ketosteroid-Δ1-dehydrogenase from Mycobacterium neoaurum JC-12.
abstract: The enzyme 3-ketosteroid-Δ(1)-dehydrogenase (KSDD), involved in steroid metabolism, catalyzes the transformation of 4-androstene-3,17-dione (AD) to androst-1,4-diene-3,17-dione (ADD) specifically. Its coding gene was obtained from Mycobacterium neoaurum JC-12 and expressed on the plasmid pMA5 in Bacillus subtilis 168. The successfully expressed KSDD was analyzed by native-PAGE. The activities of the recombinant enzyme in B. subtilis were 1.75 U/mg, which was about 5-fold that of the wild type in M. neoaurum. When using the whole-cells as catalysts, the products were analyzed by tin-layer chromatography and high-performance liquid chromatography. The recombinant B. subtilis catalyzed the biotransformation of AD to ADD in a percent conversion of 65.7% and showed about 18 folds higher than M. neoaurum JC-12. The time required for transformation of AD to ADD was about 10h by the recombinant B. subtilis, much shorter than that of the wild-type strain and other reported strains. Thus, the efficiency of ADD production could be improved immensely. For industrial applications, the recombinant B. subtilis containing KSDD provides a new pathway of producing steroid medicines.
authors.affiliation: The Key Laboratory of Industrial Biotechnology, Ministry of Education, Laboratory of Applied Microbiology and Metabolic Engineering, School of Biotechnology, Jiangnan University, Jiangsu Province, Wuxi 214122, China.
authors.forename: Wenqing
authors.initials: W
authors.lastname: Zhang
authors.forename: Minglong
authors.initials: M
authors.lastname: Shao
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Rao
authors.forename: Meijuan
authors.initials: M
authors.lastname: Xu
authors.forename: Xian
authors.initials: X
authors.lastname: Zhang
authors.forename: Taowei
authors.initials: T
authors.lastname: Yang
authors.forename: Hui
authors.initials: H
authors.lastname: Li
authors.forename: Zhenghong
authors.initials: Z
authors.lastname: Xu
mesh_terms: D000730:Androstadienes / Q000378:metabolism*; D000735:Androstenedione / Q000737:chemistry / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D055162:Biocatalysis; D015870:Gene Expression; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D009170:Nontuberculous Mycobacteria / Q000201:enzymology / Q000235:genetics*; D010088:Oxidoreductases / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA
chemical_list: D000730:Androstadienes; D011994:Recombinant Proteins; C006573:1,4-androstadiene-3,17-dione; D000735:Androstenedione; D010088:Oxidoreductases; C022656:3-oxosteroid delta(1) dehydrogenase