Figure 4.

Single-molecule reactions of the WT₆SM_ket1 pore. (A) A WT₆SM_ket1 pore was reacted with MePEG-hydroxylamine (MPHA, 1.1 kDa, 2 mM, inset) added to the trans compartment. Reaction occurred at a positive potential (+75 mV) and led to a permanent current blockade of the WT₆SM_ket1 pore. The modified pore only opened at negative applied potentials. (B) The pore was restored to an open state when 20 mM HONH₂·HCl was added to both compartments to release the PEG chain. The currents in (A) and (B) were filtered at 5 kHz and sampled at 25 kHz. For display, further digital filtering was carried out at 2 kHz with an 8-pole low-pass Bessel filter. The buffer was 1 M KCl, 50 mM Na acetate (pH 3.4). (C) Reversible oxime formation in a single synthetic pore containing a ketone (WT₆SM_ket1). Oxime formation with MPHA leads to a current drop, while reversal with HONH₂ returns the current to its initial level. The section defined by the orange bracket is magnified in panel D. (D) Negative potential (−75 mV, b) was applied during the PEG-oxime state (a), which opened the pore (residual current, I_RES = 91%). Subsequently, a positive potential (+75 mV, c) was applied, and the pore closed. The pore became fully open (violet arrow) with I_RES = 100% at negative potential (−75 mV, d), presumably when the formation of an oxime with HONH₂ led to release of the pore-bound polymer. The pore was restored to an open state at a positive potential (+75 mV, e).