Book of Abstracts: Albany 2003
June 17-21 2003
Three-dimensional Structure of a Gene Transfer Complex from Agrobacterium tumefaciens
The soil bacterium Agrobacterium tumefaciens represents the only known case of inter-kingdom DNA transfer. It infects its plant host by transfer of a linear, single-stranded DNA (T-DNA) from a tumor-inducing plasmid to the host cell genome. Expression of the encoded genes causes crown gall disease, in which tumorous growths produce compounds that only the bacterium can metabolize. Genes on the T-DNA can be manipulated or replaced, making Agrobacterium a primary vector for genetic modification of plants.
Two bacterial virulence proteins are primarily responsible for packaging of T-DNA in preparation for import to the plant cell nucleus. VirD2 binds to a sequence-specific border site on the 5' end. VirE2 binds stoichiometrically along the length of the DNA strand, without sequence specificity. A previous study of this complex using scanning transmission electron microscopy (1) showed that VirE2 packages ssDNA into a solenoid structure, and determined basic dimensional and stoichiometric parameters.
We present here a three-dimensional structure of the T-DNA-VirE2 complex by electron microscopy and image processing using a single-particle approach to helical reconstruction (2). The structure reveals a helix of ~16 nm diameter, with a hollow interior of ~7 nm diameter, 5.1 nm pitch, and 4.3 VirE2 subunits per turn. We propose that the site ssDNA lies along a continuous shelf of density that winds along the interior surface of the helix. This structure appears to be ideally suited both for protection of the ssDNA from cytoplasmic nucleases and for transport through the nuclear pore complex.
1Department of Materials and Interfaces