A new steroid-transforming strain of Mycobacterium neoaurum and cloning of 3-ketosteroid 9alpha-hydroxylase in NwIB-01.
Using enrichment procedures, five strains that can utilize soybean phytosterols as the sole carbon source were isolated from steroids-contaminated soil samples. Among the isolated strains, the strain NwIB-01 with the highest steroid degradation ability was identified as Mycobacterium neoaurum by morphological, physiological, biochemical tests and 16S rRNA sequence analysis. Meanwhile, the key enzyme gene, which was involved in steroid metabolism and encoding 395-amino acid 3-ketosteroid 9alpha-hydroxylase (KSH), was obtained from M. neoaurum NwIB-01 with the assistance of homology analysis and chromosome walking. To our best knowledge, this is the first report to the gene of key enzyme KSH from M. neoaurum. Strain NwIB-01 exhibited powerful ability of cleaving the side chain specifically from soybean phytosterols to accumulate 4-androstene-3,17-dione (AD) and 1,4-androstadiene-3,17-dione (ADD). It was showed that when cultured in 15 g/l phytosterols, the yield of ADD reached 4.23 g/l while accompanied by 1.76 g/l AD in 96-h-old culture (the molar yield of AD + ADD is 64.7%). The strain NwIB-01 can be applied as excellent phytosterols-transformation strains in potential industrial applications.
Wei
Wei
W
State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, People's Republic of China.
Fan
Shuyue
S
Wang
Fengqing
F
Wei
Dongzhi
D
D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D005798:Genes, Bacterial / Q000235:genetics; D053858:Metabolic Networks and Pathways; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000201:enzymology* / Q000235:genetics*; D010641:Phenotype; D010802:Phylogeny; D010840:Phytosterols / Q000737:chemistry / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013256:Steroids / Q000737:chemistry / Q000378:metabolism*; D013997:Time Factors
D010840:Phytosterols; D013256:Steroids; D006899:Mixed Function Oxygenases