Identification and Characterization of the LysR-Type Transcriptional Regulator HsdR for Steroid-Inducible Expression of the 3 alpha-Hydroxysteroid Dehydrogenase/Carbonyl Reductase Gene in Comamonas testosteroni

Gong, Wenjie, Xiong, Guangming and Maser, Edmund (2012) Identification and Characterization of the LysR-Type Transcriptional Regulator HsdR for Steroid-Inducible Expression of the 3 alpha-Hydroxysteroid Dehydrogenase/Carbonyl Reductase Gene in Comamonas testosteroni Applied and Environmental Microbiology, 78 (4). pp. 941-950.

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Abstract

3 alpha-Hydroxysteroid dehydrogenase/carbonyl reductase (3 alpha-HSD/CR) from Comamonas testosteroni is a key enzyme in steroid degradation in soil and water. 3 alpha-HSD/CR gene (hsdA) expression can be induced by steroids like testosterone and progester-one. Previously, we have shown that the induction of hsdA expression by steroids is a derepression where steroidal inducers bind to two repressors, RepA and RepB, thereby preventing the blocking of hsdA transcription and translation, respectively (G. Xiong and E. Maser, J. Biol. Chem. 276:9961-9970, 2001; G. Xiong, H. J. Martin, and E. Maser, J. Biol. Chem. 278:47400-47407, 2003). In the present study, a new LysR-type transcriptional factor, HsdR, for 3 alpha-HSD/CR expression in C. testosteroni has been identified. The hsdR gene is located 2.58 kb downstream from hsdA on the C. testosteroni ATCC 11996 chromosome with an orientation opposite that of hsdA. The hsdR gene was cloned and recombinant HsdR protein was produced, as was anti-HsdR polyclonal antibodies. While heterologous transformation systems revealed that HsdR activates the expression of the hsdA gene, electrophoresis mobility shift assays showed that HsdR specifically binds to the hsdA promoter region. Interestingly, the activity of HsdR is dependent on decreased repression by RepA. Furthermore, in vitro binding assays indicated that HsdR can come into contact with RNA polymerase. As expected, an hsdR knockout mutant expressed low levels of 3 alpha-HSD/CR compared to that of wild-type C. testosteroni after testosterone induction. In conclusion, HsdR is a positive transcription factor for the hsdA gene and promotes the induction of 3 alpha-HSD/CR expression in C. testosteroni.

Document Type: Article
Research affiliation: OceanRep > The Future Ocean - Cluster of Excellence
Kiel University
ISSN: 0099-2240
Projects: Future Ocean
Date Deposited: 14 May 2014 10:07
Last Modified: 14 May 2014 10:07
URI: http://eprints.uni-kiel.de/id/eprint/23993

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