Molecular dynamics simulations of ion permeation and gating in potassium channels
by Andrei Mironenko
Date of Examination:2025-01-28
Date of issue:2025-03-13
Advisor:Dr. Wojciech Kopec
Referee:Prof. Dr. Andreas Janshoff
Referee:Prof. Dr. Markus Zweckstetter
Referee:Prof. Dr. Luis A. Pardo
Referee:Prof. Dr. Ricardo Mata
Referee:Prof. Dr. Marcus Müller
Persistent Address:
http://dx.doi.org/10.53846/goediss-11143
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Abstract
English
Potassium (K+) channels are transmembrane ion channels that conduct K+ ions with exceptional selectivity and permeation rates. K+ channels participate in a plethora of important cellular processes, including termination of action potentials in excitable cells. This thesis addresses the two main phenomena associated with K+ channels - mechanisms of ion permeation and gating of K+ channel activity - using molecular dynamics (MD) simulations. In studies of K+ channels, the use of both experimental and computational methods has significantly contributed to our understanding of K+ channel structure and function. Among these methods, MD simulations allow studying the processes relevant to K+ channels at time and space resolutions often inaccessible to other approaches. In the present thesis, first the physical basis and practical aspects of using MD simulations are described, along with strengths and limitations of the method. Then, I focus on K+ channels, and discuss the two most common models of K+ permeation, soft and direct knock-on, along with the available experimental and computational data addressing the two models. I conclude the introduction to the thesis by addressing the gating mechanisms - regulation of the equilibrium between conductive and non-conductive states of a channel, together with the associated changes in the channel conformation - described for various K+ channel families. In this thesis, ion permeation mechanisms in K+ channels are studied specifically on the example of a model K+ channel, KcsA, and its selectivity filter (SF) mutants. The SF is the narrowest part of the ion permeation pathway, largely conserved between different K+ channel families. Generally, most structures of K+ channels exhibit a direct knock-on-like ion configuration in the SF, characterized by the absence of water molecules and presence of close ion-ion contacts. Certain point mutations in the SF, however, allow to isolate soft knock-on-like configurations, where neighboring ions are separated by water molecules. This, coupled with the specific effects these mutations have on the ion occupancy in non-adjacent K+ binding sites, was proposed to favor the soft knock-on mechanism in the wild type channels as well. Here, we use MD simulations with an applied electric field to study ion permeation in the WT and mutant KcsA. By testing various combinations of force fields, mutations and voltages, we show not only that ion permeation mechanisms are fundamentally different in mutant and WT channels, but also that the complete loss of the ion solvation shell upon entry and close ion-ion contacts (direct knock-on mechanism) in the SF are crucial to establish the hallmark high conductance and ion selectivity in K+ channels. The second project is focused on gating in BK channels. BK channels are activated by membrane depolarization and binding of Ca2+, which is facilitated by a complex structure featuring the pore, voltage- and Ca2+-sensing domains. Despite several CryoEM structures capturing the full-length BK in activating and deactivating conditions, the mechanism by which ion permeation is stopped in the closed state of the channel is unclear, as the pore in deactivating conditions lacks any physical constrictions often present in other K+ channels. By performing atomistic simulations of the full-length BK, we suggest that occupancy of the pore by lipids defines the closed, non-conductive state of the channel. In the corresponding part of the thesis, I describe the mechanisms by which lipid entry into the pore occurred, as well as the associated effects on the channel’s conductance. Then, I address the effect of membrane composition on BK activity. It was shown that, generally, negatively charged lipids increase both the open probability and single-channel current in BK. In good agreement with the experimental data, our coarse-grained and atomistic MD simulations suggest a multi-modal mechanism, in which the negative charge on the lipid headgroups leads to reduced lipid entry, increased local K+ concentration in the pore and increased conformational stability of the open-state channel. Finally, in ‘Conclusions and outlook’ I summarize the findings of the two projects, and discuss the approaches to further validate our conclusions, and expand upon the corresponding research questions.
Keywords: Potassium channels; Molecular dynamics simulations
