VirD4 is an ATPase that selects proteins to be secreted in most T4SSs. We have shown that VirD4 from Xac recognizes conserved domains called XVIPCD found at the C-termini of X-Tfes (Alegria et al., 2005). We are interested in understanding the structure of the VirD4-XVIPCD complex, how the complex is transfered to the rest of the T4SS machinery and the nature of the role played by ATP binding and/or hydrolysis. Since the transfer of toxins by the X. citri T4SS kills recipient E. coli cells, we will produce a X. citri strain in which all of the genes coding for X-Tfes (XVIPS) have been knocked out. This non-killer strain, with an otherwise functional T4SS, will be transformed with plasmids expressing specific X-Tfes. If this is successful, this assay could be used to quantitatively measure transfer modified X-Tfes with mutations in specific recognition motifs. This X. citri strain will also be used to screen mutations in other T4SS components to map the structural requirements for secretion.
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