Poster Presentation The 3rd Prato Conference on Pore Forming Proteins 2015

Electrophysiological studies of the interaction of polymers with pores formed by bacterial toxins (1) provide a window on single molecule interaction with proteins in real time, (2) report on the behavior of macromolecules in confinement and (3) enable label-free single molecule sensing technologies. It has been shown previously that the electrical current through pores formed by the staphylococcal toxin alpha-hemolysin (aHL) in a lipid bilayer is blocked for periods of hundreds of microseconds to milliseconds by polyethylene glycol (PEG) oligomers (degree of polymerization approx. 10-60). Notably, this block showed monomeric sensitivity on PEG mass, allowing the construction of mass spectra from the residual current values. Here, we show that the electrical current through an aerolysin pore (AeL) from Aeromonas hydrophila is also blocked by PEG but with important differences in the voltage-dependence of the interaction kinetics. While PEG blocking events of aHL show maximum residence times at a stemside-positive transmembrane voltage of about +40 mV and minimum residence times at stemside-negative voltages, blocks of AeL are so short as to be not resolvable (Low pass filter cut off = 20 kHz) at stemside-positive voltages but increase in duration with increasing values of stemside-negative voltage up to -200 mV. Importantly, the interaction also shows monomer sensitivity to PEG mass, and at >-100 mV stemside-negative voltage long residence times and high driving force combine to provide particularly precise determination of residual current, resulting in high peak-to-floor ratio mass spectra. These findings may potentially be understood as a consequence of the relatively high content of negative charges of the AeL pore compared to aHL and suggest that comparative studies of polymer interactions with different pore proteins are important in elucidating the underlying physicochemical mechanisms.