Molecular characterization ofthevirDoperon fromAgrobacterium tumefaciens

TheAgrobacterium tumefaciens Ti plasmidvirulence(vir)region containsat leastsixtranscriptional unitsrequiredfortheefficient transferofT-DNAto theplantgenome(virA,B, C, D, E, andG). We have reportedthattwoproteinsencodedby the5'portionof thevirDoperonare requiredfora site-specific endonuclease activitythatnicksthedirect repeatswhichflanktheT-DNA. We havepresentedthenucleotidesequence forthisportionof theoperon.Thenucleotidesequenceof theremainder of thevirDoperonessential forvirulencehasnowbeendetermined.Two additional openreadingframesencodeproteinsof 21.3and75.8kilodaltons (kd).Translational fusionsbetweenvirD2,virD3,andvirD4proteinsand trpEproducedfusionproteinsof thesizepredictedfromthenucleotide sequencedata. We haveusedantiseradirectedagainstthetrpE-virD2 fusionproteinto detectbothnativevirD2proteinanda virD2-lacZfusion proteinin crudeextractsfromAgrobacterium.