Biology subjects

Kopec, W.

Publications and source records attributed to Kopec, W..

3 recordsLinked to original sources

bioRxiv2023

Selectivity filter mutations shift ion permeation mechanism in potassium channels

Potassium (K+) channels combine high conductance with high ion selectivity. To explain this efficiency, two molecular mechanisms have been proposed. The direct knock-on mechanism is defined by water-free K+ permeation and formation of direct ion-ion contacts in the highly conserved selectivity filter (SF). The soft knock-on mechanism involves co-permeation of water and separation of K+ by water molecules. With the aim to distinguish between these mechanisms, crystal structures of the KcsA channel with mutations in two SF residues - G77 and T75 - were published, where the arrangements of K+ ions and water display canonical soft knock-on configurations. These data were interpreted as evidence of the soft knock-on mechanism in wild-type channels (C. Tilegenova, et al., Structure, function, and ion-binding properties of a K+ channel stabilized in the 2,4-ion-bound configuration. Proceedings of the National Academy of Sciences 116, 16829-16834 (2019)). Here, we test this interpretation using molecular dynamics simulations of KcsA and its mutants. We show that, while a strictly water-free direct knock-on permeation is observed in the wild-type, conformational changes induced by these mutations lead to distinct ion permeation mechanisms, characterized by co-permeation of K+ and water. These mechanisms are characterized by reduced conductance and impaired potassium selectivity, supporting the importance of full dehydration of potassium ions for the hallmark high conductance and selectivity of K+ channels. In general, we present a case where mutations introduced at the critical points of the permeation pathway in an ion channel drastically change its permeation mechanism in a non-intuitive manner. Significance statementPotassium (K+) channels conduct K+ with high permeation rates and ion selectivity. An ongoing debate in the field has been focused on the molecular mechanisms underlying this remarkable efficiency. Here, we performed molecular dynamics simulations of two selectivity filter mutants of a model K+ channel to investigate this question. These mutations led to a substantial decrease in conductance and ion selectivity, while accompanied by a shift from water-free K+ permeation to co-permeation of water and K+. Our findings not only provide a fundamental example of how single point mutations in the selectivity filter can alter the ion permeation mechanism, but also reinforce the notion that water exclusion underlies the remarkable efficiency of K+ channels.

biophysics↗

bioRxiv2022

Periodic boundaries in Molecular Dynamics simulations: why do we need salt?

Molecular dynamics (MD) simulations are usually performed by employing periodic boundary conditions (PBC). While this treatment of simulation system removes the necessity to treat the interactions with an otherwise artificial boundary, PBC also introduces additional constraints that need to be carefully considered for a robust and reliable simulation. Some of the issues pertaining to PBC are well explored and can be remedied by choosing a large enough unit cell, or by applying corrections to the generated trajectories. In current work, we study another artifact which cannot be alleviated by changing the box size. The artifact occurs due to the PBC imposed constraints affecting systems with permanent uncompensated dipoles, which is of particular relevance for lipid membrane simulations. Such dipoles often arise in many biologically-relevant setups, in particular those involving asymmetric lipid bilayers. The artifact manifests itself as an electric field formation in the simulation box which is counteracted by redistribution of mobile charge carriers (ions) and/or ordering of water dipoles. In the absence of ions, the artifact may cause strong water ordering, affecting thermodynamics of the studied system. This observation reveals a conceptually interesting effect of using explicit salt in MD simulations: ions help removing the unwanted periodicity-induced artifact occurring due to uncompensated electric dipoles. Therefore, we recommend adding mobile ions in molecular simulations whenever possible, and call for caution when simulating systems that require low salt concentration (or no salt at all), for example ion channel inactivation promoting conditions. In general, our findings are relevant for molecular simulations of any systems that contain uncompensated dipoles, that might occur more often than previously thought.

biophysics↗

bioRxiv2022

Driving Forces underlying Selectivity Filter Gating in the MthK Potassium Channel

K+ channel activity can be limited by C-type inactivation, which is likely initiated in part by dissociation of K+ ions from the selectivity filter, and modulated by side chains surrounding the selectivity filter. Whereas crystallographic and computational studies have linked inactivation to a collapsed selectivity filter conformation in the KcsA channel, the structural basis for selectivity filter gating in other K+ channels has been less clear. Here, we combined electrophysiological recordings with molecular dynamics based, in silico electrophysiology simulations, to study selectivity filter gating in the model potassium channel MthK and its V55E mutant (analogous to KcsA E71) in the pore-helix. Experimentally, we find that MthK V55E has a lower open probability than the WT channel, due to decreased stability of the open state, as well as a lower unitary conductance. Simulations account for both aspects of these observations on the atomistic scale, showing that ion permeation in V55E is altered by two distinct orientations of the E55 side chain. In the vertical orientation of E55, in which E55 forms a hydrogen bond with D64 (as observed with KcsA WT channels), the filter displays reduced conductance compared to MthK WT. In contrast, with horizontal orientation, K+ conductance is closer to MthK WT; however the selectivity filter stability in the conducting conformation is lowered, and the filter more readily transitions to the inactivated conformation. Surprisingly, these transitions of MthK WT and V55E channels to the non-conducting (inactivated) state observed in simulations are associated with a widening selectivity filter, unlike its narrowing seen in KcsA, and reminisce the recent structures of stably-inactivated, voltage-gated potassium channels: Shaker W434F and Kv1.2 W362F mutants, as well as WT Kv1.3 channels.

biophysics↗