PilV Adhesins of Plasmid R64 Thin Pili Specifically Bind to the Lipopolysaccharides of Recipient Cells

IncI1 plasmid R64 encodes type IV pili or thin pili, which contain PilV adhesins. The C-terminal segments of PilV adhesins are exchanged into seven types by shufflon multiple DNA inversion. PilV adhesins determine recipient specificity in R64 liquid matings through the recognition of lipopolysaccharides (LPSs) on the surface of recipient cells. Using various waa mutants of Escherichia coli R1 as recipient cells, liquid mating experiments suggest that PilVA adhesin recognizes the GlcNAc(β1–3)Glc moiety of E. coli R1 type LPS. The direct binding of PilV adhesins to LPSs of the recipient bacterial strains was demonstrated using filter overlay assays. The specificity of PilV-LPS binding is in close agreement with the recipient specificity determined by R64 liquid matings. The C-terminal segments of PilVA, PilVC, PilVC′, and PilVD′ adhesins were expressed as fusion proteins with glutathione- S-transferase (GST). GST-A, GST-C, GST-C′, and GST-D′ proteins bound to their respective LPSs with the specificities identical with those determined in the R64 liquid matings, indicating that the C-terminal segments of PilV adhesins bind to specific moieties of LPS molecules.