Analysis of VirB Proteins of A. Tumefaciens Required for T-DNA Transfer

根癌农杆菌的毒力菌株通过将T-DNA从其驻留的钛质粒转移到植物细胞中，从而启动植物上的肿瘤。细菌与植物细胞之间的有效相互作用需要细菌与植物细胞的适当结合，并在细菌中表达一系列毒力(Vir)基因。虽然已知几种病毒蛋白的功能，但T-DNA通过两组细胞壁和细胞膜运输的机制尚不清楚。最近的数据支持DNA转移的机制类似于细菌接合的假说，这表明DNA转移机制，如大肠杆菌F质粒编码的接合小孔，参与了DNA转移。VIRB操纵子是编码这种孔中蛋白质的合理候选者。VIRB蛋白在T-DNA从细菌到植物细胞的转移中的可能作用将通过体外诱变和重组DNA技术、蛋白质的免疫表征和免疫定位以及确定蛋白质是否参与与细菌膜或DNA的物理相互作用的生物化学方法来研究。目前，植物生物技术很大程度上依赖于以农杆菌为基础的基因转移。一些植物很容易被这种载体转化，而另一些则不能。因此，阐明DNA在该系统中的转移机制对植物基因工程具有重要意义。

Ward Virulent strains of Agrobacterium tumefaciens initiate tumors on plants by transferring the T-DNA from its resident Ti plasmid into plant cells. A productive interaction between bacterium and plant cell requires the appropriate attachment of the bacterium to the plant cell, and expression of a series of virulence (vir) genes in the bacterium. While the functions of several vir proteins are known, the mechanism by which the T-DNA is transported across two sets of cell walls and membranes is not known. Recent data supports the hypothesis that the mechanism of DNA transfer is analogous to that of bacterial conjugation, suggesting that a DNA transfer apparatus, such as the conjugal pore encoded by the F plasmid of E. coli, is involved. The virB operon is the logical candidate to encode the proteins of such a pore. The possible role of virB proteins in the transfer of T- DNA from the bacterium to the plant cell will be investigated using in vitro mutagenesis and recombinant DNA techniques, immunocharacterization and immunolocalization of the proteins, and biochemical approaches to determine whether or not the proteins are involved in physical interactions with bacterial membranes or DNA. Currently, plant biotechnology is heavily dependent on gene transfer based on Agrobacterium. Some plants are readily transformable by this vector, while others are not. Thus, elucidating the mechanism of DNA transfer in this system will be of great importance to plant genetic engineering.