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Identification of promoters recognized by RNA polymerase containing Mycobacterium tuberculosis stress-response sigma factor σF

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Abstract

A previously optimized Escherichia coli two-plasmid system was used to identify Mycobacterium tuberculosis promoters recognized by RNA polymerase containing the M. tuberculosis stress response sigma factor σF. The method allowed the identification of five new σF-dependent promoters. Transcriptional start points of the promoters were determined by high-resolution S1-nuclease mapping using RNA prepared from E. coli containing the two-plasmid system. The promoters were confirmed by an in vitro transcription assay. The Mycobacterium smegmatis and Mycobacterium tuberculosis core RNA polymerases, after complementation with σF, were able to recognize all the five promoters. All the promoters contained sequences highly similar to the sequence of the previously identified M. tuberculosis σF-dependent promoter, usfXp1. Comparison of the promoters revealed a σF consensus sequence GtTtga-N14–18–GGGTAT. The σF-dependent promoters may govern expression of genes encoding a transcription regulator homologous to the response regulators of bacterial two-component signal transduction systems and proteins with unknown function.

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Acknowledgments

We are grateful to Prof. P. J. Brennan and Dr. J. T. Belisle (Colorado State University, Fort Collins, CO, USA; NIH, NIAID Contract N01 AI-75320) for chromosomal DNA from M. tuberculosis H37Rv, and to Dr. M.K.Winson (University of Nottingham, UK) for plasmid pSB40. We would like to thank Dr. S. Rodrigue (University of Sherbrooke, Canada) for plasmids pSR52, pJF09, and pJF10. We are grateful also to Eva Kutejova and Beata Sevcikova for help in isolation of M. smegmatis RNA polymerase. This work was supported by a Grant 2/6010/26 from Slovak Academy of Sciences and sponsored by the US—Slovak Science and Technology joint fund under project number 032/2001.

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Authors and Affiliations

  1. Institute of Molecular Biology, Center of Excellence for Molecular Medicine, Slovak Academy of Sciences, Dubravska cesta 21, 845 51, Bratislava, Slovak Republic

    Dagmar Homerova, Katarina Surdova & Jan Kormanec

  2. Department of Biochemistry, Faculty of Natural Sciences, Comenius University, 842 15, Bratislava, Slovak Republic

    Katarina Mikusova

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  1. Dagmar Homerova
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  2. Katarina Surdova
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  3. Katarina Mikusova
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  4. Jan Kormanec
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Correspondence to Jan Kormanec.

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Homerova, D., Surdova, K., Mikusova, K. et al. Identification of promoters recognized by RNA polymerase containing Mycobacterium tuberculosis stress-response sigma factor σF . Arch Microbiol 187, 185–197 (2007). https://doi.org/10.1007/s00203-006-0185-6

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  • DOI: https://doi.org/10.1007/s00203-006-0185-6

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