The virA protein of Agrobacterium tumefaciens is autophosphorylated and is essential for vir gene regulation

• Published in: Journal of Bacteriology Vol. 172; no. 2; pp. 525 - 530
• Main Authors: Jin, S.G. (University of Washington, Seattle, WA), Roitsch, T, Ankenbauer, R.G, Gordon, M.P, Nester, E.W
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.02.1990
• Subjects: 550201 - Biochemistry- Tracer Techniques; AGROBACTERIUM; AMINO ACIDS; Amino Acids - analysis; ATP; AZOLES; BACTERIA; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; Bacteriology; Base Sequence; BASIC BIOLOGICAL SCIENCES; BETA DECAY RADIOISOTOPES; BETA-MINUS DECAY RADIOISOTOPES; Biological and medical sciences; CARBOXYLIC ACIDS; CELL CONSTITUENTS; CHEMICAL REACTIONS; Cloning, Molecular; DAYS LIVING RADIOISOTOPES; DISEASES; ESCHERICHIA COLI; Escherichia coli - genetics; FOSFORILACION; Fundamental and applied biological sciences. Psychology; GENE; Gene Expression Regulation, Bacterial; GENE REGULATION; GENES; Genes, Bacterial; HETEROCYCLIC ACIDS; HETEROCYCLIC COMPOUNDS; HISTIDINE; IMIDAZOLES; ISOTOPES; LIGHT NUCLEI; Microbiology; MICROORGANISMS; Molecular Sequence Data; NEOPLASMS; NUCLEI; NUCLEOTIDES; ODD-ODD NUCLEI; Oligonucleotide Probes; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANIC NITROGEN COMPOUNDS; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; PATHOGENS; PHOSPHORUS 32; PHOSPHORUS ISOTOPES; PHOSPHORYLATION; PLANTS; PLASMIDE; PLASMIDIOS; PLASMIDS; PROTEINAS; PROTEINE; RADIOISOTOPES; Rhizobium - genetics; Rhizobium - metabolism; Rhizobium - pathogenicity; TRANSCRIPTION; Virulence; Virulence Factors; Autophosphorylation; Phosphorylation; Radiolabelling; Gel electrophoresis; Virulence; Site directed mutagenesis; Agrobacterium tumefaciens; Phosphorus; Phytopathogen; Gene expression; Proteins; Regulation(control); Histidine; Gene; Bacteria; Rhizobiaceae
• ISSN: 0021-9193; 1098-5530; 1067-8832
• DOI: 10.1128/jb.172.2.525-530.1990

The virA and virG gene products are required for the regulation of the vir regulon on the tumor-inducing (Ti) plasmid of Agrobacterium tumefaciens. VirA is a membrane-associated protein which is homologous to the sensor molecules of other two-component regulatory systems. We overproduced truncated VirA proteins in Escherichia coli by deleting different lengths of the 5'-coding region of the virA gene and placing these genes under lacZ control. These proteins were purified from polyacrylamide gels and renatured. The renatured proteins became radiolabeled when they were incubated with [-gamma- 32P]ATP but not with [gamma-32P]GTP or [alpha-32P]ATP, which suggests an ATP gamma-phosphate-specific autophosphorylation. The smallest VirA protein, which retained only the C-terminal half of the protein, gave the strongest autophosphorylation signal, which demonstrates that the C-terminal domain has the autophosphorylation site. The phosphorylated amino acid was identified as phosphohistidine, and a highly conserved histidine was found in all of the VirA homologs. When this histidine was changed to glutamine, which cannot be phosphorylated, the resulting VirA protein lost both its ability to autophosphorylate and its biological function as a vir gene regulator. Results of this study indicate that VirA autophosphorylation is required for the induction of the vir regulon and subsequent tumor induction on plants by A. tumefaciens