Mutational analysis of the VirG protein, a transcriptional activator of Agrobacterium tumefaciens virulence genes.
The VirG protein of Agrobacterium tumefaciens is required in conjunction with the VirA protein for transcriptional activation of the virulence (vir) genes in response to plant phenolic compounds. These proteins are members of a family of two component regulatory systems. vir genes are activated via a cascade of phosphorylation reactions involving a specific aspartic acid residue of the VirG protein. We have conducted a mutational analysis of the VirG protein. By mutating conserved and nonconserved aspartic acid residues in the N-terminal domain, we demonstrated that two of three conserved aspartic acid residues located in two different regions are important for the phosphorylation of VirG by VirA phosphate. A third conserved N-terminal region was also shown to be critical for the biological function of VirG as a transcriptional activator. The identification of phosphorylatable but biologically inactive mutated VirG proteins suggests that not only phosphorylation but also a conformational change is necessary for its activity. We further demonstrated that phosphorylation is not required for sequence-specific binding to a vir gene regulatory sequence (vir box) and that the C-terminal domain is sufficient for DNA binding. The data support the model of a two-domain structure for the VirG protein and demonstrate that the sequence homologies to other two-component regulatory systems reflect both functional and structural homologies.
Department of Microbiology, University of Washington, Seattle 98195.
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Roitsch
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Wang
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Jin
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Nester
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012231:Rhizobium / Q000235:genetics* / Q000472:pathogenicity; D012689:Sequence Homology, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics*; D014774:Virulence / Q000235:genetics
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D014157:Transcription Factors; C058483:virG protein, Shigella flexneri