A mutation in the receiver domain of the Agrobacterium tumefaciens transcriptional regulator VirG increases its affinity for operator DNA.
We fused the wild-type Agrobacterium tumefaciens virG gene and the constitutive virGN54D allele to the malE gene of Escherichia coli, and studied the binding of MBP-VirG fusions to the autoregulated virG promoter. MBP-VirGN54D protein bound this promoter with 10-fold higher affinity than MBP-VirG, and bound to vir box I with eightfold higher affinity than to vir box III. Disruption of vir box III did not alter the affinity for vir box I, suggesting a lack of cooperativity between these sites. We provide evidence that protein bound at a single vir box may have a higher oligomeric state than non-bound protein, and that a DNA distortion adjacent to vir box I may occur during activation.
Han
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DC
Section of Microbiology, Cornell University, Ithaca, New York 14853.
Winans
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SC
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D001426:Bacterial Proteins; D001483:Base Sequence; D004269:DNA, Bacterial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005809:Genes, Regulator; D008969:Molecular Sequence Data; D009875:Operator Regions, Genetic; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C058483:virG protein, Shigella flexneri