Solving the Gating Mechanism of the Mitochondrial β-Barrel Metabolite Channel VDAC

Abstract

The Voltage-Dependent Anion Channel (VDAC) is a β-barrel protein at the mitochondrial outer membrane (MOM) serving as the major pathway for ions and respiratory metabolites between mitochondria and the cytosol. Under applied voltage, VDAC transitions from an anionic “open” state, allowing the passive diffusion of neutral and anionic metabolites, to multiple low-conducting “closed” states with reduced anion selectivity. Importantly, VDAC closed states are virtually impermeable to ATP. A potential implication of these closed states in regulating metabolite traffic across the MOM has triggered vigorous investigations on their structural nature. Still, the mechanism behind VDAC closure remains enigmatic. Here, we study VDAC1 voltage-gating by using ion-channel electrophysiology guided by ANTON2 multi-microsecond Molecular Dynamics (MD) simulations. MD simulations highlight the particular role played by conformational dynamics of a “trigger” region, which is especially impacted by voltage polarity or solution acidification. We found that a basic residue in this region is key for initiating gating transitions at neutral pH, and its mutation to the oppositely charged residue hinders channel closure and reverses selectivity. We further explore VDAC voltage-gating by combining selectivity and pH-titration experiments with Markov State Model analysis on the motions of charged residues obtained from MD simulations. We propose a VDAC gating mechanism in which all charged residues are part of an ensemble where the motion of the single basic residue at the trigger region acts as a seed coordinating distinct dynamic rearrangements of the salt-bridge complex network. Thus, VDAC closure does not imply significant conformational changes of the β-barrel. This is strikingly different from the gating mechanism of most highly ion-selective channels and thus establishes a new paradigm of β-barrel channel gating that does not involve the translocation of a voltage sensor or a major change in pore geometry.