Three-dimensional reconstruction of Agrobacterium VirE2 protein with single-stranded DNA.
Agrobacterium tumefaciens infects plant cells by a unique mechanism involving an interkingdom genetic transfer. A single-stranded DNA substrate is transported across the two cell walls along with the bacterial virulence proteins VirD2 and VirE2. A single VirD2 molecule covalently binds to the 5'-end of the single-stranded DNA, while the VirE2 protein binds stoichiometrically along the length of the DNA, without sequence specificity. An earlier transmission/scanning transmission electron microscopy study indicated a solenoidal ("telephone coil") organization of the VirE2-DNA complex. Here we report a three-dimensional reconstruction of this complex using electron microscopy and single-particle image-processing methods. We find a hollow helical structure of 15.7-nm outer diameter, with a helical rise of 51.5 nm and 4.25 VirE2 proteins/turn. The inner face of the protein units contains a continuous wall and an inward protruding shelf. These structures appear to accommodate the DNA binding. Such a quaternary arrangement naturally sequesters the DNA from cytoplasmic nucleases and suggests a mechanism for its nuclear import by decoration with host cell factors. Coexisting with the helices, we also found VirE2 tetrameric ring structures. A two-dimensional average of the latter confirms the major features of the three-dimensional reconstruction.
Abu-Arish
Asmahan
A
Department of Materials and Interfaces, Weizmann Institute of Science, Rehovot 76100, Israel.
Frenkiel-Krispin
Daphna
D
Fricke
Tobin
T
Tzfira
Tzvi
T
Citovsky
Vitaly
V
Wolf
Sharon Grayer
SG
Elbaum
Michael
M
D021581:Active Transport, Cell Nucleus; D001426:Bacterial Proteins; D004277:DNA, Single-Stranded / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry*; D021621:Imaging, Three-Dimensional; D007473:Ion Channels / Q000737:chemistry*; D008854:Microscopy, Electron; D009690:Nucleic Acid Conformation; D012231:Rhizobium / Q000737:chemistry*
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D007473:Ion Channels; C051583:virE2 protein, Agrobacterium