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PMID:2394678

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Citation

Jin, SG, Prusti, RK, Roitsch, T, Ankenbauer, RG and Nester, EW (1990) Phosphorylation of the VirG protein of Agrobacterium tumefaciens by the autophosphorylated VirA protein: essential role in biological activity of VirG. J. Bacteriol. 172:4945-50

Abstract

Agrobacterium tumefaciens virulence genes are induced by plant signals through the VirA-VirG two-component regulatory system. The VirA protein is a membrane-spanning sensor molecule that possesses an autophosphorylating activity, and the VirG protein is a sequence-specific DNA-binding protein. In this report, we demonstrate that the VirG protein is phosphorylated by the VirA protein and that the phosphate is directly transferred from the phosphorylated VirA molecule (phosphohistidine) to the VirG protein. The chemical stability of the phospho-VirG bond suggested that the VirG protein was phosphorylated at the aspartate and/or glutamate residue. The phosphorylated VirG protein was reduced with tritiated sodium borohydride and subjected to proteolytic digestion with the Achromobacter protease I enzyme. The resulting peptide fragments were separated by C8 reversed-phase high-pressure liquid chromatography, and the tritium-labeled peptide was sequenced. Amino acid sequence data showed that the aspartate residue at position 52 was the only site phosphorylated. Changing this aspartate into asparagine resulted in a nonphosphorylatable and biologically nonfunctional gene product. As a control, a randomly chosen aspartate was changed into an asparagine (position 72), and no effect on its phosphorylation or biological activity was observed. Unlike its homologs, including CheA-CheY, EnvZ-OmpR, and NtrB-NtrC, the phospho-VirG molecule was very stable in vitro. The possible implications of these observations and the function of VirG phosphorylation in vir gene activation are discussed.

Links

PubMed PMC213149

Keywords

Aspartic Acid; Bacterial Proteins/isolation & purification; Bacterial Proteins/metabolism; Chromatography, High Pressure Liquid; DNA-Binding Proteins; Mutation; Phosphoproteins/isolation & purification; Phosphorylation; Plasmids; Rhizobium/genetics; Rhizobium/metabolism; Transcription Factors; Virulence Factors

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

AGRT5:VIRA

GO:0004673: protein histidine kinase activity

ECO:0000314:

F

Figure 1 shows that VirA has the ability to phosphorylate VirG. Figure 2 shows that the phosphorylation is through a phosphotransfer reaction. Figure 5 shows that the phosphotransfer is to a specific aspartate residue in VirG, a reaction typical of a protein histidine kinase.

complete

AGRT5:VIRA

GO:0016301: kinase activity

ECO:0000314:

F

Figure 1 shows that VirA has the ability to phosphorylate VirG. Figure 2 shows that the phosphorylation is through a phosphotransfer reaction. Figure 5 shows that the phosphotransfer is to a specific aspartate residue in VirG, a reaction typical of a protein histidine kinase.

complete

AGRT5:VIRA

GO:0016310: phosphorylation

ECO:0000314:

P

Figure 1 shows that VirA has the ability to phosphorylate VirG. Figure 2 shows that the phosphorylation is through a phosphotransfer reaction. Figure 5 shows that the phosphotransfer is to a specific aspartate residue in VirG, a reaction typical of a protein histidine kinase.

complete


See also

References

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