Inter-aminoacid residues electrostatic interactions contribute to the conformational stability of peptides and proteins, influence their folding pathways, and are critically important to a multitude of problems in biology including the onset of misfolding diseases. By varying the pH and ionic strength, the inter-aminoacid residues electrostatic interactions of histidine-containing, β-hairpin-like peptides, alter their folding behavior, and we studied this through quantifying at the uni-molecular level the frequency, dwell-times of translocation events and amplitude of blockades associated to interactions between such peptides and the α-hemolysin (α-HL) protein. Acidic buffers were shown to dram... More
Inter-aminoacid residues electrostatic interactions contribute to the conformational stability of peptides and proteins, influence their folding pathways, and are critically important to a multitude of problems in biology including the onset of misfolding diseases. By varying the pH and ionic strength, the inter-aminoacid residues electrostatic interactions of histidine-containing, β-hairpin-like peptides, alter their folding behavior, and we studied this through quantifying at the uni-molecular level the frequency, dwell-times of translocation events and amplitude of blockades associated to interactions between such peptides and the α-hemolysin (α-HL) protein. Acidic buffers were shown to dramatically decrease the rate of peptide capture by the α-HL protein, through the interplay of enthalpic and entropic contributions brought about on the free energy barrier which controls the peptides-α-HL association rate. We found that in acidic buffers, the amplitude of the blockage induced by a α-HL's β-barrel-residing peptide is smaller than the value seen at neutral pH, and this supports our interpretation of the pH-induced change in the conformation of the peptide, which behaves as a less-stable hairpin at acidic pH values that obstructs to a lesser extent the protein pore. This is also confirmed by the fact that the dissociation rate of such model peptide from the α-HL's β-barrel is higher at acidic, as compared to neutral pH values. Experiments performed in low-salt buffers revealed the dramatic decrease of the peptide capture rate by the α-HL protein, most likely caused by the increase in the radius of counterions cloud around the peptide, thus hindering peptide's partition into the β-barrel. We posit that the reduced electrostatic screening in low-salt buffers leads to a decrease in peptides effective cross-sectional area and their mobility within the α-HL pore, caused by the peptide chain stretching augmentation via increased inter-residues electrostatic interactions, and histidines protonation in acidic, low-salt buffers, bolsters this effect.
