Fig. 5: Evidence in support of a structural deformation responsible for gating.

a, Sketch illustrating both states of the model. The pore exhibits mechanical bistability induced by depletion forces: when the pore diameter fluctuates below a certain threshold, ions are expelled from the pore, causing it to collapse partially. This effect is promoted by the dissociation of counterions from lumen charges under an applied voltage. b, Closed-state probability plotted against the phase of the voltage for aerolysin wt and four representative mutants obtained from experiments (left) and the biophysical model (right) (see Supplementary Section 2 for details). The model accurately predicts whether the given mutant is a positive, a negative or a double gater. It also predicts that the E254A mutant gates stronger than aerolysin wt. c, Scatter plot of the pH response and all 26 aerolysin mutants’ closing rates versus rectification factor β. Pores that show low gating or gating at both polarities are represented twice. For pores that show gating only in the negative or positive quadrant, only one value is shown. The error bars show ±1 standard deviation of the mean over the number of experimental replicas n, and each symbol represents a different mutant (Supplementary Fig. 11). Dotted line: theoretical prediction based on models of rectification and gating (Supplementary Section 2). d, Cryo-EM images of E254A–E258A mutant (left) and aerolysin wt (right). Boxes delimited with dashed lines show corresponding 2D classes of the two pore populations; the box size is 290 angstrom. e, Stack of a mutant pore (green) and a mutant prepore (dark green) (left) and alignment of the structures of wt (light green) and the E254A–E258A (9GXJ) mutant (green) of a fully formed pore (right). f, Closing rates of aerolysin wt for different membrane thicknesses (left) and of m2MspA and m2MspA cross-linked with glutaral and methanal (right) (1 M KCl, 10 mM phosphate, pH 6.2, 0.1 Hz). The error bars show ±1 standard deviation of the mean over the number of experimental replicas n.