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Journal of Bacteriology, October 2003, p. 6215-6219, Vol. 185, No. 20
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.20.6215-6219.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Nucleotide-Dependent Conformational Changes in the {sigma}54-Dependent Activator DctD

Ying-Kai Wang,1,{dagger} Sungdae Park,2,{ddagger} B. Tracy Nixon,2 and Timothy R. Hoover1*

Department of Microbiology, University of Georgia, Athens, Georgia,1 Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania2

Received 25 April 2003/ Accepted 29 July 2003

Activators of {sigma}54-RNA polymerase holoenzyme couple ATP hydrolysis to formation of an open promoter complex. DctD{Delta}1-142, a truncated and constitutively active form of the {sigma}54-dependent activator DctD from Sinorhizobium meliloti, displayed an altered DNase I footprint at its binding site located upstream of the dctA promoter in the presence of ATP. The altered footprint was not observed for a mutant protein with a substitution at or near the putative arginine finger, a conserved arginine residue thought to contact the nucleotide. These data suggest that structural changes in DctD{Delta}1-142 during ATP hydrolysis can be detected by alterations in the DNase I footprint of the protein and may be communicated by interactions between bound nucleotide and the arginine finger. In addition, kinetic data for changes in fluorescence energy transfer upon binding of 2'(3')-O-(N-methylanthraniloyl)-ATP (Mant-ATP) to DctD{Delta}1-142 and DctD suggested that these proteins undergo multiple conformational changes following ATP binding.

* Corresponding author. Mailing address: 527 Biological Science Building, University of Georgia, Athens, GA 30602. Phone: (706) 542-2675. Fax: (706) 542-2674. E-mail: trhoover{at}arches.uga.edu.

{dagger} Present address: Bristol-Myers Squibb, Pharmaceutical Research Institute, Wallingford, CT 06492.

{ddagger} Present address: Department of Biochemistry and Molecular Biology, Medical College of Ohio, Toledo, OH 43614-5804.

Journal of Bacteriology, October 2003, p. 6215-6219, Vol. 185, No. 20
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.20.6215-6219.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Burrows, P. C., Joly, N., Nixon, B. T., Buck, M. (2009). Comparative analysis of activator-E{sigma}54 complexes formed with nucleotide-metal fluoride analogues. Nucleic Acids Res 37: 5138-5150 [Abstract] [Full Text]  
  • De Carlo, S., Chen, B., Hoover, T. R., Kondrashkina, E., Nogales, E., Nixon, B. T. (2006). The structural basis for regulated assembly and function of the transcriptional activator NtrC. Genes Dev. 20: 1485-1495 [Abstract] [Full Text]  
  • Xu, H., Gu, B., Nixon, B. T., Hoover, T. R. (2004). Purification and Characterization of the AAA+ Domain of Sinorhizobium meliloti DctD, a {sigma}54-Dependent Transcriptional Activator. J. Bacteriol. 186: 3499-3507 [Abstract] [Full Text]  


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