Alterations within the activation domain of the sigma 54-dependent activator DctD that prevent transcriptional activation.
AUTOR(ES)
Wang, Y K
RESUMO
Rhizobium meliloti DctD (C4-dicarboxylate transport protein D) is a transcriptional activator that catalyzes the ATP-dependent isomerization of closed complexes between sigma 54-RNA polymerase holoenzyme and the dctA promoter to open complexes. Following random mutagenesis of dctD, 55 independent mutant forms of DctD that failed to activate transcription from a dctA'-'lacZ reporter gene in Escherichia coli were selected, and the amino acid substitutions were determined for these mutant proteins. Amino acid substitutions were distributed throughout the central domain of the protein, the domain responsible for transcription activation, but most of the substitutions occurred within three highly conserved regions of the protein. Selected mutant proteins were purified, and their activities were studied in vitro. All of the purified mutant proteins appeared to have normal DNA-binding activity and interacted with sigma 54 and core RNA polymerase, as determined from protein crosslinking assays. Proteins with amino acid substitutions in a region spanning amino acid positions 222 to 225 retained their ATPase activities, whereas proteins with substitutions in other regions had little or no ATPase activity. Taken together, these data suggest that the region that encompasses amino acid residues 222 through 225 probably functions in coupling the energy released from ATP hydrolysis to open complex formation rather than as a major determinant for binding to RNA polymerase.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=179471Documentos Relacionados
- Negative regulation of sigma 54-dependent dctA expression by the transcriptional activator DctD.
- Rhizobium meliloti and Rhizobium leguminosarum dctD gene products bind to tandem sites in an activation sequence located upstream of sigma 54-dependent dctA promoters.
- Nucleotide-Dependent Conformational Changes in the σ54-Dependent Activator DctD
- Purification and Characterization of the AAA+ Domain of Sinorhizobium meliloti DctD, a σ54-Dependent Transcriptional Activator
- Mutational Analysis of the Phosphate-Binding Loop of Rhizobium meliloti DctD, a ς54-Dependent Activator