Fig 7.

Effects of VirB11 substrate discrimination mutations on DNA transfer and VirB10 activation. Upper, transfer of T-DNA and IncQ plasmid pML122 to VirD4, native and mutant forms of VirB11, and VirB6 as assessed with the TrIP assay. Strains: A348 (WT strain) without and with pML122 (IncQ plasmid); PC1011(pML122) (ΔvirB11 mutant carrying pML122) without (−) or with plasmids expressing B11 (native VirB11) or i4 mutant proteins (the i4 mutation is inserted after the residue listed). S, supernatant from the IP reaction; IP, immunoprecipitated material recovered with anti-VirD4, -VirB11, or -VirB6 antibodies (αD4, αB11, αB6). For PC1011 strains producing native or mutant forms of VirB11, amplicons from the immunoprecipitate (IP) fractions are shown. Ct, control, Ti plasmid amplicon; IncQ, pML122 plasmid amplicon; T-DNA, T-DNA amplicon. Lower, A348(pML122) and PC1011(pML122) strains were assayed for accumulation of B10 (native VirB10) and B10* (VirB10*) upon protease treatment of spheroplasts by immunostaining with the anti-VirB10 antibodies. T-DNA, T-DNA transfer to plants as monitored by virulence (+, virulent; −, avirulent); IncQ, transfer of IncQ plasmid pML122 to agrobacterial recipients.