This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Abo-Amer, A. E. Articles by Andrews, S. C. Search for Related Content PubMed PubMed Citation Articles by Abo-Amer, A. E. Articles by Andrews, S. C.

 Previous Article  |  Next Article 

Journal of Bacteriology, March 2004, p. 1879-1889, Vol. 186, No. 6
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.6.1879-1889.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

DNA Interaction and Phosphotransfer of the C₄-Dicarboxylate- Responsive DcuS-DcuR Two-Component Regulatory System from Escherichia coli

Aly E. Abo-Amer,¹ Jonathan Munn,¹ Kerry Jackson,¹ Murat Aktas,^1, Paul Golby,^1, David J. Kelly,² and Simon C. Andrews^1*

The School of Animal and Microbial Sciences, University of Reading, Whiteknights, Reading RG6AJ,¹ Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield S10 2TN, United Kingdom²

Received 18 September 2003/ Accepted 17 November 2003

The DcuS-DcuR system of Escherichia coli is a two-component sensor-regulator that controls gene expression in response to external C₄-dicarboxylates and citrate. The DcuS protein is particularly interesting since it contains two PAS domains, namely a periplasmic C₄-dicarboxylate-sensing PAS domain (PASp) and a cytosolic PAS domain (PASc) of uncertain function. For a study of the role of the PASc domain, three different fragments of DcuS were overproduced and examined: they were PASc-kinase, PASc, and kinase. The two kinase-domain-containing fragments were autophosphorylated by [-³²P]ATP. The rate was not affected by fumarate or succinate, supporting the role of the PASp domain in C₄-dicarboxylate sensing. Both of the phosphorylated DcuS constructs were able to rapidly pass their phosphoryl groups to DcuR, and after phosphorylation, DcuR dephosphorylated rapidly. No prosthetic group or significant quantity of metal was found associated with either of the PASc-containing proteins. The DNA-binding specificity of DcuR was studied by use of the pure protein. It was found to be converted from a monomer to a dimer upon acetylphosphate treatment, and native polyacrylamide gel electrophoresis suggested that it can oligomerize. DcuR specifically bound to the promoters of the three known DcuSR-regulated genes (dctA, dcuB, and frdA), with apparent K_Ds of 6 to 32 µM for untreated DcuR and 1 to 2 µM for the acetylphosphate-treated form. The binding sites were located by DNase I footprinting, allowing a putative DcuR-binding motif [tandemly repeated (T/A)(A/T)(T/C)(A/T)AA sequences] to be identified. The DcuR-binding sites of the dcuB, dctA, and frdA genes were located 27, 94, and 86 bp, respectively, upstream of the corresponding +1 sites, and a new promoter was identified for dcuB that responds to DcuR.

Present address: Heinrich Heine University of Duesseldorf, Duesseldorf, Germany.

Present address: VLA, Weybridge, Surrey KT15 3NB, United Kingdom.

This article has been cited by other articles:

• Scheu, P., Sdorra, S., Liao, Y.-F., Wegner, M., Basche, T., Unden, G., Erker, W. (2008). Polar accumulation of the metabolic sensory histidine kinases DcuS and CitA in Escherichia coli. Microbiology 154: 2463-2472 [Abstract] [Full Text]  
• Shulami, S., Zaide, G., Zolotnitsky, G., Langut, Y., Feld, G., Sonenshein, A. L., Shoham, Y. (2007). A Two-Component System Regulates the Expression of an ABC Transporter for Xylo-Oligosaccharides in Geobacillus stearothermophilus. Appl. Environ. Microbiol. 73: 874-884 [Abstract] [Full Text]  
• Depardieu, F., Podglajen, I., Leclercq, R., Collatz, E., Courvalin, P. (2007). Modes and Modulations of Antibiotic Resistance Gene Expression. Clin. Microbiol. Rev. 20: 79-114 [Abstract] [Full Text]  
• Wei, J.-R., Tsai, Y.-H., Soo, P.-C., Horng, Y.-T., Hsieh, S.-C., Ho, S.-W., Lai, H.-C. (2005). Biochemical Characterization of RssA-RssB, a Two-Component Signal Transduction System Regulating Swarming Behavior in Serratia marcescens. J. Bacteriol. 187: 5683-5690 [Abstract] [Full Text]  
• Satomura, T., Shimura, D., Asai, K., Sadaie, Y., Hirooka, K., Fujita, Y. (2005). Enhancement of Glutamine Utilization in Bacillus subtilis through the GlnK-GlnL Two-Component Regulatory System. J. Bacteriol. 187: 4813-4821 [Abstract] [Full Text]  
• Goh, E.-B., Bledsoe, P. J., Chen, L.-L., Gyaneshwar, P., Stewart, V., Igo, M. M. (2005). Hierarchical Control of Anaerobic Gene Expression in Escherichia coli K-12: the Nitrate-Responsive NarX-NarL Regulatory System Represses Synthesis of the Fumarate-Responsive DcuS-DcuR Regulatory System. J. Bacteriol. 187: 4890-4899 [Abstract] [Full Text]  
• Ramos, J. L., Martinez-Bueno, M., Molina-Henares, A. J., Teran, W., Watanabe, K., Zhang, X., Gallegos, M. T., Brennan, R., Tobes, R. (2005). The TetR Family of Transcriptional Repressors. Microbiol. Mol. Biol. Rev. 69: 326-356 [Abstract] [Full Text]  
• Kneuper, H., Janausch, I. G., Vijayan, V., Zweckstetter, M., Bock, V., Griesinger, C., Unden, G. (2005). The Nature of the Stimulus and of the Fumarate Binding Site of the Fumarate Sensor DcuS of Escherichia coli. J. Biol. Chem. 280: 20596-20603 [Abstract] [Full Text]  
• Yamamoto, K., Hirao, K., Oshima, T., Aiba, H., Utsumi, R., Ishihama, A. (2005). Functional Characterization in Vitro of All Two-component Signal Transduction Systems from Escherichia coli. J. Biol. Chem. 280: 1448-1456 [Abstract] [Full Text]  
• Janausch, I. G., Garcia-Moreno, I., Lehnen, D., Zeuner, Y., Unden, G. (2004). Phosphorylation and DNA binding of the regulator DcuR of the fumarate-responsive two-component system DcuSR of Escherichia coli. Microbiology 150: 877-883 [Abstract] [Full Text]