The Origin of Coupled Chloride and Proton Transport in a Cl^–/H⁺ Antiporter

Abstract

The ClC family of transmembrane proteins functions throughout nature to control the transport of Cl– ions across biological membranes. ClC-ec1 from Escherichia coli is an antiporter, coupling the transport of Cl– and H+ ions in opposite directions and driven by the concentration gradients of the ions. Despite keen interest in this protein, the molecular mechanism of the Cl–/H+ coupling has not been fully elucidated. Here, we have used multiscale simulation to help identify the essential mechanism of the Cl–/H+ coupling. We find that the highest barrier for proton transport (PT) from the intra- to extracellular solution is attributable to a chemical reaction, the deprotonation of glutamic acid 148 (E148). This barrier is significantly reduced by the binding of Cl– in the “central” site (Cl–cen), which displaces E148 and thereby facilitates its deprotonation. Conversely, in the absence of Cl–cen E148 favors the “down” conformation, which results in a much higher cumulative rotation and deprotonation barrier that effectively blocks PT to the extracellular solution. Thus, the rotation of E148 plays a critical role in defining the Cl–/H+ coupling. As a control, we have also simulated PT in the ClC-ec1 E148A mutant to further understand the role of this residue. Replacement with a non-protonatable residue greatly increases the free energy barrier for PT from E203 to the extracellular solution, explaining the experimental result that PT in E148A is blocked whether or not Cl–cen is present. The results presented here suggest both how a chemical reaction can control the rate of PT and also how it can provide a mechanism for a coupling of the two ion transport processes.

Multiscale Kinetic Modeling Reveals an Ensemble of Cl–/H+ Exchange Pathways in ClC-ec1 Antiporter

“…We note that this definition is slightly different than used in our previous work. 35 In that work, we defined a “down” state with a Cl – at S cen for when E148 was rotated to interact with water molecules leading into the pore region, noting that the side chain could not reach as far into the pore as when S cen was not occupied by a chloride ion. That position in the pore corresponds to the lower basin in the region here defined as “up” in panel Figure 5 B.…”

Dynamical model of the CLC-2 ion channel reveals conformational changes associated with selectivity-filter gating

“…Experimental biophysicists have shown that these mechanisms are complex, involving interdependent influences of transmembrane voltage, [Cl − ], and [H + ] [8,[32][33][34][35][36]. To begin to resolve these complexities and reveal the dynamic, atomistic details of the CLC gating machine, it will be critical to pair structural studies with high-resolution conformational modeling [37][38][39][40][41][42][43][44][45][46].…”

Multiscale Simulation Reveals Passive Proton Transport Through SERCA on the Microsecond Timescale

“…The MS-RMD models are developed from and calibrated against QM/MM data by utilizing a force matching algorithm in a "machine learning" type methodology. The MS-RMD approach can efficiently and accurately simulate explicit PT in proteins, as mediated by water molecules and amino acids while including Grotthuss proton shuttling, see, e.g., refs (34)(35)(36)(41)(42)(43)(44)(45).…”