The mechanical behavior of individual P pili of uropathogenic Escherichia coli has been investigated using optical tweezers. P pili, whose main part constitutes the PapA rod, composed of approximately 10(3) PapA subunits in a helical arrangement, are distributed over the bacterial surface and mediate adhesion to host cells. They are particularly important in the pathogenesis of E. coli colonizing the upper urinary tract and kidneys. A biological model system has been established for in situ measurements of the forces that occur during mechanical stretching of pili. A mathematical model of the force-versus-elongation behavior of an individual pilus has been developed. Three elongation regions of pili were identified. In region I, P pili stretch elastically, up to a relative elongation of 16 +/- 3%. The product of elasticity modulus and area of a P pilus, EA, was assessed to 154 +/- 20 pN (n=6). In region II, the quaternary structure of the PapA rod unfolds under a constant force of 27 +/- 2 pN (n approximately 100) by a sequential breaking of the interactions between adjacent layers of PapA subunits. This unfolding can elongate the pilus up to 7 +/- 2 times. In region III, pili elongate in a nonlinear manner as a result of stretching until the bond ruptures.

P pili are protein filaments expressed by uropathogenic Escherichia coli that mediate binding to glycolipids on epithelial cell surfaces, which is a prerequisite for bacterial infection. When a bacterium, attached to a cell surface, is exposed to external forces, the pili, which are composed of B10 3 PapA protein subunits arranged in a helical conformation, can elongate by unfolding to a linear conformation. This property is considered important for the ability of a bacterium to withstand shear forces caused by urine flow. It has hitherto been assumed that this elongation is plastic, thus constituting a permanent conformational deformation. We demonstrate, using optical tweezers, that this is not the case; the unfolding of the helical structure to a linear conformation is fully reversible. It is surmised that this reversibility helps the bacteria regain close contact to the host cells after exposure to significant shear forces, which is believed to facilitate their colonization.

SUMMARY The BabA adhesin mediates high-affinity binding of Helicobacter pylori to the ABO blood-group antigen-glycosylated gastric mucosa. Here we show that BabA is acid responsive but binding is restored by pH neutralization. Acid responsiveness differs among strains; often correlates with different intragastric regions; changes during chronic infection and disease progression; and depends on pH sensor sequences in BabA and on pH reversible formation of high-affinity binding BabA multimers. We propose that BabA’s extraordinary reversible acid-responsiveness enables tight mucosal bacterial adherence while at the same time allowing an effective escape from epithelial cells and mucus that are shed into the acidic bactericidal lumen and that bio-selection and changes in BabA binding properties through mutations and recombination with babA-related genes are selected by differences among individuals and by changes in gastric acidity over time. These processes generate diverse H. pylori subpopulations, and BabA’s adaptive evolution contributes importantly to H. pylori persistence and to overt gastric disease.

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P-pili of uropathogenic Escherichia coli mediate the attachment to epithelial cells in the human urinary tract and kidney and therefore play an important role in infection. A better understanding of this mechanism could help to prevent bacteria from spreading but also provides interesting insights into molecular mechanics for future nanotech applications. The helical rod design of P-pili provides an efficient design to withstand hydrodynamic shear forces. The adhesive PapG unit at the distal end of the P-pilus forms a specific bond with the glycolipid Galabiose. This bond has a potential width Deltax = 0.7 +/- 0.15 nm and a dissociation rate K (Off) = 8.0.10(-4) +/- 5.0.10(-4) s(-1). It withstands a force of approximately 49 pN under physiological conditions. Additionally, we analyzed the behavior of unstacking and restacking of the P-pilus with dynamic force spectroscopy at velocities between 200 and 7,000 nm/s. Up to a critical extension of 66% of the totally stretched P-pilus, un/re-stacking was found to be fully reversible at velocities up to 200 nm/s. If the P-pilus is stretched beyond this critical extension a characteristic hysteresis appears upon restacking. This hysteresis originates from a nucleation process comparable to a first-order phase transition in an undercooled liquid. Analysis of the measurement data suggests that 20 PapA monomers are involved in the formation of a nucleation kernel.