Structure and function of conjugative pili: monoclonal antibodies as probes for structural variants of F pili

The lac-tra operon fusion plasmid pTG801 contains the known F plasmid DNA transfer (tra) genes required by Escherichia coli to elaborate functional F pili (T. Grossman and P. M. Silverman, J. Bacteriol. 171:650-656, 1989). Here, we show that these pili are actually structural variants of normal F pili and that the F plasmid must contain additional genes that affect pilus structure and function. We confirmed a previous report that two monoclonal antibodies that recognize epitopes at and near the amino terminus of F pilin do not decorate the sides of normal F pili, as determined by immunogold electron microscopy. However, both antibodies laterally decorated pTG801 pili. The epitope for one of the antibodies has been shown to include the amino-terminal acetyl group of F pilin, which must therefore also be present on pTG801 pilin. Normal antibody staining was restored to pTG801 pili when cells contained, in addition to pTG801, the compatible plasmid pRS31, which must therefore include at least one gene affecting F-pilus structure. One candidate, traD, was excluded as the sole such gene, since traD+ derivatives of a pTG801 strain still elaborated pili that could be laterally decorated with antibody. Moreover, although traD alone restored RNA bacteriophage R17 infectivity to pTG801 cells, as expected, it did not mimic pRS31 in restoring to pTG801 pili other characteristics of normal F pili. We conclude that pRS31 contains as yet uncharacterized genes required for elaboration of structurally normal F pili. Finally, we identified vesicular material, especially abundant in cultures of pTG801 transformants, that stained heavily with the anti-F-pilin monoclonal antibodies. This material may reflect the inner membrane pool of F pilin.