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Journal of Bacteriology, September 2004, p. 6277-6285, Vol. 186, No. 18
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.18.6277-6285.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Amino Acid Contacts between Sigma 70 Domain 4 and the Transcription Activators RhaS and RhaR

Jason R. Wickstrum and Susan M. Egan*

Department of Molecular Biosciences, University of Kansas, Lawrence, Kansas

Received 17 May 2004/ Accepted 24 June 2004

The RhaS and RhaR proteins are transcription activators that respond to the availability of L-rhamnose and activate transcription of the operons in the Escherichia coli L-rhamnose catabolic regulon. RhaR activates transcription of rhaSR, and RhaS activates transcription of the operon that encodes the L-rhamnose catabolic enzymes, rhaBAD, as well as the operon that encodes the L-rhamnose transport protein, rhaT. RhaS is 30% identical to RhaR at the amino acid level, and both are members of the AraC/XylS family of transcription activators. The RhaS and RhaR binding sites overlap the –35 hexamers of the promoters they regulate, suggesting they may contact the {sigma}70 subunit of RNA polymerase as part of their mechanisms of transcription activation. In support of this hypothesis, our lab previously identified an interaction between RhaS residue D241 and {sigma}70 residue R599. In the present study, we first identified two positively charged amino acids in {sigma}70, K593 and R599, and three negatively charged amino acids in RhaR, D276, E284, and D285, that were important for RhaR-mediated transcription activation of the rhaSR operon. Using a genetic loss-of-contact approach we have obtained evidence for a specific contact between RhaR D276 and {sigma}70 R599. Finally, previous results from our lab separately showed that RhaS D250A and {sigma}70 K593A were defective at the rhaBAD promoter. Our genetic loss-of-contact analysis of these residues indicates that they identify a second site of contact between RhaS and {sigma}70.

* Corresponding author. Mailing address: Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045. Phone: (785) 864-4294. Fax: (785) 864-5294. E-mail: sme{at}ku.edu.

Journal of Bacteriology, September 2004, p. 6277-6285, Vol. 186, No. 18
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.18.6277-6285.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Dominguez-Cuevas, P., Marin, P., Busby, S., Ramos, J. L., Marques, S. (2008). Roles of Effectors in XylS-Dependent Transcription Activation: Intramolecular Domain Derepression and DNA Binding. J. Bacteriol. 190: 3118-3128 [Abstract] [Full Text]  
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  • Kolin, A., Jevtic, V., Swint-Kruse, L., Egan, S. M. (2007). Linker Regions of the RhaS and RhaR Proteins. J. Bacteriol. 189: 269-271 [Abstract] [Full Text]  
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