The VirE1VirE2 complex of Agrobacterium tumefaciens interacts with single-stranded DNA and forms channels.
The VirE2 protein is crucial for the transfer of single-stranded DNA (ssDNA) from Agrobacterium tumefaciens to the nucleus of the plant host cell because of its ssDNA binding activity, assistance in nuclear import and putative ssDNA channel activity. The native form of VirE2 in Agrobacterium's cytoplasm is in complex with its specific chaperone, VirE1. Here, we describe the ability of the VirE1VirE2 complex to both bind ssDNA and form channels. The affinity of the VirE1VirE2 complex for ssDNA is slightly reduced compared with VirE2, but the kinetics of binding to ssDNA are unaffected by the presence of VirE1. Upon binding of VirE1VirE2 to ssDNA, similar helical structures to those reported for the VirE2-ssDNA complex were observed by electron microscopy. The VirE1VirE2 complex can release VirE1 once the VirE2-ssDNA complexes assembled. VirE2 exhibits a low affinity for small unilamellar vesicles composed of bacterial lipids and a high affinity for lipid vesicles containing sterols and sphingolipids, typical components of animal and plant membranes. In contrast, the VirE1VirE2 complex associated similarly with all kind of lipids. Finally, black lipid membrane experiments revealed the ability of the VirE1VirE2 complex to form channels. However, the majority of the channels displayed a conductance that was a third of the conductance of VirE2 channels. Our results demonstrate that the binding of VirE1 to VirE2 does not inhibit VirE2 functions and that the effector-chaperone complex is multifunctional.
Duckely
Myriam
M
M.E. Müller Institute for Structural Biology, Biozentrum, University of Basel, Basel, CH-4056, Switzerland.
Oomen
Clasien
C
Axthelm
Fabian
F
Van Gelder
Patrick
P
Waksman
Gabriel
G
Engel
Andreas
A
D016960:Agrobacterium tumefaciens / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D007473:Ion Channels / Q000378:metabolism*; D050356:Lipid Metabolism; D046911:Macromolecular Substances / Q000378:metabolism; D008566:Membranes / Q000378:metabolism; D008854:Microscopy, Electron; D018832:Molecular Chaperones / Q000378:metabolism*
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D007473:Ion Channels; D046911:Macromolecular Substances; D018832:Molecular Chaperones; C051582:virE1 protein, Agrobacterium tumefaciens; C051583:virE2 protein, Agrobacterium