Membrane Position Dependency of the pKa and Conductivity of the Protein Ion Channel

Abstract

The dependency of current-voltage characteristics of the α-hemolysin channel on the channel position within the membrane was studied using Poisson-Nernst-Planck theory of ion conductivity with soft repulsion between mobile ions and protein atoms (SP-PNP). The presence of the membrane environment also influences the protonation state of the residues at the boundary of the water-lipid interface. In this work, we predict that Asp and Lys residues at the protein rim change their protonation state upon penetration to the lipid environment. Free energies of protein insertion in the membrane for different penetration depths were estimated using the Poisson-Boltzmann/solvent-accessible surface area (PB/SASA) model. The results show that rectification and reversal potentials are very sensitive to the relative position of channel in the membrane, which in turn contributes to alternative protonation states of lipid-penetrating ionizable groups. The prediction of channel position based on the matching of calculated rectification with experimentally determined rectification is in good agreement with recent neutron reflection experiments. Based on the results, we conclude that α-hemolysin membrane position is determined by a combination of factors and not only by the pattern of the surface hydrophobicity as is typically assumed.

Keywords: Current–voltage characteristics; Ion channels; Ion conductivity; Poisson–Nernst–Plank theory; pKa in membrane; α-Hemolysin.

Figure 1

The structure and positioning of the AHL channel in the membrane. Two extreme positions are shown: the purple slab corresponds to deep insertion of AHL and the green slab corresponds to shallow insertion. The molecular surface of the channel represents ion accessible surface; the surface is colored by the residue types: white is used for hydrophobic residues, green - for polar residues, blue and red - for positively and negatively charged residues respectively. The shown system boundaries are not up to scale. This figure was prepared with the VMD program[53].

Figure 2

Dependencies of the electrostatic potential along the channel pore on penetration depth (d) of AHL to the membrane. Contributions of protein, membrane and water are shown. The protonation state is as used for I-V calculations. Continuum electrostatic potentials are also compared to the electrostatic potential from all-atom MD simulations calculated by Aksimentiev and Schulten [22].

Figure 3

Total current (A) and rectification (B) versus position of AHL in the membrane. The position of the channel corresponding for the position when the center of channel’s hydrophobic belt center coincides with membrane center is shown by dark gray vertical solid line; the experimental position [14] of the channel is shown by dark gray vertical dashed line. The results of simulations are compared to experimental results of Menestrina [54], Merzlyak et al. [55], Krasilnikov et al. [56] and Walker et al. [57].

Figure 4

Dependency of the reversal potential (A) and the selectivity (B) on the AHL penetration depths. The position of the channel corresponding to the position when the center of channel’s hydrophobic belt center coincides with membrane center is shown by gray vertical line; the experimental position[14] of the channel is shown by gray vertical dashed line. The results of simulations are compared to experimental results: Menestrina[54] and Gu et al.[58]. The selectivity as current ratio (B) is shown for 0.1 M KCl.

Figure 5

A - Change in free energy of insertion on the different levels of penetration of AHL to the membrane. B – Same, focused on the minimum at 30 Å.