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Applied and Environmental Microbiology, April 2003, p. 2139-2152, Vol. 69, No. 4
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.4.2139-2152.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Gene Encoding the Hydrolase for the Product of the meta-Cleavage Reaction in Testosterone Degradation by Comamonas testosteroni

Masae Horinouchi,1* Toshiaki Hayashi,1 Hiroyuki Koshino,1 Takako Yamamoto,1 and Toshiaki Kudo1,2,3

RIKEN,1 JST, Wako-shi, Saitama 351-0198,2 Science of Biological Supermolecular Systems, Graduate School of Integrated Science, Yokohama City University, Suehiro, Tsurumi-ku, Yokohama 230-0045, Japan3

Received 17 October 2002/ Accepted 20 January 2003

In a previous study we isolated the meta-cleavage enzyme gene, tesB, that encodes an enzyme that carries out a meta-cleavage reaction in the breakdown of testosterone by Comamonas testeroni TA441 (M. Horinouchi et al., Microbiology 147:3367-3375, 2001). Here we report the isolation of a gene, tesD, that encodes a hydrolase which acts on the product of the meta-cleavage reaction. We isolated tesD by using a Tn5 mutant of TA441 that showed limited growth on testosterone. TesD exhibited ca. 40% identity in amino acid sequence with BphDs, known hydrolases of biphenyl degradation in Pseudomonas spp. The TesD-disrupted mutant showed limited growth on testosterone, and the culture shows an intense yellow color. High-pressure liquid chromatography analysis of the culture of TesD-disrupted mutant incubated with testosterone detected five major intermediate compounds, one of which, showing yellow color under neutral conditions, was considered to be the product of the meta-cleavage reaction. The methylation product was analyzed and identified as methyl-4,5-9,10-diseco-3-methoxy-5,9,17-trioxoandrosta-1(10),2-dien-4-oate, indicating that the substrate of TesD in testosterone degradation is 4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-dien-4-oic acid. 4,5-9,10-Diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-dien-4-oic acid was transformed by Escherichia coli-expressed TesD. Downstream of tesD, we identified tesE, F, and G, which encode for enzymes that degrade one of the products of 4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-dien-4-oic acid converted by TesD.

* Corresponding author. Mailing address: RIKEN, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan. Phone: 81-48-467-9545. Fax: 81-48-462-4672. E-mail: masae{at}postman.riken.go.jp.

Applied and Environmental Microbiology, April 2003, p. 2139-2152, Vol. 69, No. 4
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.4.2139-2152.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Gohler, A., Xiong, G., Paulsen, S., Trentmann, G., Maser, E. (2008). Testosterone-inducible Regulator Is a Kinase That Drives Steroid Sensing and Metabolism in Comamonas testosteroni. J. Biol. Chem. 283: 17380-17390 [Abstract] [Full Text]  
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