Abstract
The gene encoding the putative reductase component (KshB) of 3-ketosteroid 9α-hydroxylase was cloned from Rhodococcus equi USA-18, a cholesterol oxidase-producing strain formerly named Arthrobacter simplex USA-18, by PCR according to consensus amino acid motifs of several bacterial KshB subunits. Deletion of the gene in R. equi USA-18 by a PCR-targeted gene disruption method resulted in a mutant strain that could accumulate up to 0.58 mg/ml 1,4-androstadiene-3,17-dione (ADD) in the culture medium when 0.2% cholesterol was used as the carbon source, indicating the involvement of the deleted enzyme in 9α-hydroxylation of steroids. In addition, this mutant also accumulated 3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid (Δ1,4-BNC). Because both ADD and Δ1,4-BNC are important intermediates for the synthesis of steroid drugs, this mutant derived from R. equi USA-18 may deserve further investigation for its application potential.
Highlights
Steroid drugs, including androgens, anabolic steroids, estrogens and corticosteroids, have been widely used for a variety of health purposes
Isolation of putative KshA and KshB genes from A. simplex USA-18 To clone KshA genes from A. simplex USA-18, the amino acid sequences of several bacterial Rieske [2Fe-2S] terminal oxygenases, including those isolated from R. jostii RHA1 (YP_704482), M. smegmatis (YP_890151), Burkholderia cenocepacia J2315 (YP_002234232), Ralstonia eutropha JMP134 (YP_295786) and Comamonas testosteroni KF-1 (WP_003057373), were aligned
The upstream and downstream regions of S1A2 were obtained by inverse PCR and DNA walking, and an S1A2containing open reading frame (ORF) of 1155 nucleotides was identified
Summary
Steroid drugs, including androgens, anabolic steroids, estrogens and corticosteroids, have been widely used for a variety of health purposes. They are produced via chemical synthesis and/or biotransformation using steroid catabolic intermediates, such as 4-androstene3,17-dione (AD) and 1,4-androstadiene-3,17-dione (ADD), as starting materials [1]. Bacterial strains belonging to Mycobacterium, Rhodococcus, Nocardia and Arthrobacter genera are known for their ability to degrade a range of naturally occurring steroids. The catabolic pathways for sterols have been proposed based on the results of a large body of biochemical and genomic studies [2,3,4].
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
