Digging into Lipid Membrane Permeation for Cardiac Ion Channel Blocker d-Sotalol with All-Atom Simulations.

Interactions of drug molecules with lipid membranes play crucial role in their accessibility of cellular targets and can be an important predictor of their therapeutic and safety profiles. Very little is known about spatial localization of various drugs in the lipid bilayers, their active form (ionization state) or translocation rates and therefore potency to bind to different sites in membrane proteins. All-atom molecular simulations may help to map drug partitioning kinetics and thermodynamics, thus providing in-depth assessment of drug lipophilicity. As a proof of principle, we evaluated extensively lipid membrane partitioning of d-sotalol, well-known blocker of a cardiac potassium channel Kv11.1 encoded by the hERG gene, with reported substantial proclivity for arrhythmogenesis. We developed the positively charged (cationic) and neutral d-sotalol models, compatible with the biomolecular CHARMM force field, and subjected them to all-atom molecular dynamics (MD) simulations of drug partitioning through hydrated lipid membranes, aiming to elucidate thermodynamics and kinetics of their translocation and thus putative propensities for hydrophobic and aqueous hERG access. We found that only a neutral form of d-sotalol accumulates in the membrane interior and can move across the bilayer within millisecond time scale, and can be relevant to a lipophilic channel access. The computed water-membrane partitioning coefficient for this form is in good agreement with experiment. There is a large energetic barrier for a cationic form of the drug, dominant in water, to cross the membrane, resulting in slow membrane translocation kinetics. However, this form of the drug can be important for an aqueous access pathway through the intracellular gate of hERG. This route will likely occur after a neutral form of a drug crosses the membrane and subsequently re-protonates. Our study serves to demonstrate a first step toward a framework for multi-scale in silico safety pharmacology, and identifies some of the challenges that lie therein

Model of ionic currents through microtubule nanopores and the lumen

It has been suggested that microtubules and other cytoskeletal filaments may act as electrical transmission lines. An electrical circuit model of the microtubule is constructed incorporating features of its cylindrical structure with nanopores in its walls. This model is used to study how ionic conductance along the lumen is affected by flux through the nanopores when an external potential is applied across its two ends. Based on the results of Brownian dynamics simulations, the nanopores were found to have asymmetric inner and outer conductances, manifested as nonlinear IV curves. Our simulations indicate that a combination of this asymmetry and an internal voltage source arising from the motion of the C-terminal tails causes a net current to be pumped across the microtubule wall and propagate down the microtubule through the lumen. This effect is demonstrated to enhance and add directly to the longitudinal current through the lumen resulting from an external voltage source, and could be significant in amplifying low-intensity endogenous currents within the cellular environment or as a nano-bioelectronic device.Comment: 43 pages, 6 figures, revised versio

Magic Angle Spinning Nuclear Magnetic Resonance Characterization of Voltage-Dependent Anion Channel Gating in Two-Dimensional Lipid Crystalline Bilayers

National Institutes of Health (U.S.) (EB001960)National Institutes of Health (EB002026

Magic Angle Spinning Nuclear Magnetic Resonance Characterization of Voltage-Dependent Anion Channel Gating in Two-Dimensional Lipid Crystalline Bilayers

The N-terminus of the voltage-dependent anion channel (VDAC) has been proposed to contain the mechanistically important gating helices that modulate channel opening and closing. In this study, we utilize magic angle spinning nuclear magnetic resonance (MAS NMR) to determine the location and structure of the N-terminus for functional channels in lipid bilayers by measuring long-range 13C–13C distances between residues in the N-terminus and other domains of VDAC reconstituted into DMPC lipid bilayers. Our structural studies show that the distance between A14 Cβ in the N-terminal helix and S193 Cβ is ∼4–6 Å. Furthermore, VDAC phosphorylation by a mitochondrial kinase at residue S193 has been claimed to delay mitochondrial cell death by causing a conformational change that closes the channel, and a VDAC-Ser193Glu mutant has been reported to show properties very similar to those of phosphorylated VDAC in a cellular context. We expressed VDAC-S193E and reconstituted it into DMPC lipid bilayers. Two-dimensional 13C–13C correlation experiments showed chemical shift perturbations for residues located in the N-terminus, indicating possible structural perturbations to that region. However, electrophysiological data recorded on VDAC-S193E showed that channel characteristics were identical to those of wild type samples, indicating that phosphorylation of S193 does not directly affect channel gating. The combination of NMR and electrophysiological results allows us to discuss the validity of proposed gating models

Bases of Bacterial Sodium Channel Selectivity Among Organic Cations

Hille’s (1971) seminal study of organic cation selectivity of eukaryotic voltage-gated sodium channels showed a sharp size cut-off for ion permeation, such that no ion possessing a methyl group was permeant. Using the prokaryotic channel, NaChBac, we found some similarity and two peculiar differences in the selectivity profiles for small polyatomic cations. First, we identified a diverse group of minimally permeant cations for wildtype NaChBac, ranging in sizes from ammonium to guanidinium and tetramethylammonium; and second, for both ammonium and hydrazinium, the charge-conserving selectivity filter mutation (E191D) yielded substantial increases in relative permeability (PX/PNa). The relative permeabilities varied inversely with relative Kd calculated from 1D Potential of Mean Force profiles (PMFs) for the single cations traversing the channel. Several of the cations bound more strongly than Na+, and hence appear to act as blockers, as well as charge carriers. Consistent with experimental observations, the E191D mutation had little impact on Na+ binding to the selectivity filter, but disrupted the binding of ammonium and hydrazinium, consequently facilitating ion permeation across the NaChBac-like filter. We concluded that for prokaryotic sodium channels, a fine balance among filter size, binding affinity, occupancy, and flexibility seems to contribute to observed functional differences