Different roles for aspartates and glutamates for cation permeation in bacterial sodium channels

Carlo Guardiani,Olena A Fedorenko,Igor A Khovanov,Stephen K Roberts

doi:10.1016/j.bbamem.2018.11.011

Abstract

A key driving force for ion channel selectivity is represented by the negative charge of the Selectivity Filter carried by aspartate (D) and glutamate (E) residues. However, the structural effects and specific properties of D and E residues have not been extensively studied. In order to investigate this issue we studied the mutants of NaChBac channel with all possible combinations of D and E in the charged rings in position 191 and 192. Electrophysiological measurements showed significant Ca2+ currents only when position 191 was occupied by E. Equilibrium Molecular Dynamics simulations revealed the existence of two binding sites, corresponding to the charged rings and another one, more internal, at the level of L190. The simulations showed that the ion in the innermost site can interact with the residue in position 191 only when this is glutamate. Based on the MD simulations, we suggest that a D in position 191 leads to a high affinity Ca2+ block site resulting from a significant drop in the free energy of binding for an ion moving between the binding sites; in contrast, the free energy change is more gradual when an E residue occupies position 191, resulting in Ca2+ permeability. This scenario is consistent with the model of ion channel selectivity through stepwise changes in binding affinity proposed by Dang and McCleskey. Our study also highlights the importance of the structure of the selectivity filter which should contribute to the development of more detailed physical models for ion channel selectivity.

Highlights

Voltage-gated sodium and calcium channels (NaV and CaV channels, respectively) are important mediators of many physiological functions, including propagation of electrical signal, cell excitability, gene transcription, secretion and synaptic transmission, plasticity and muscle contraction [1,2]
Data are presented as means ± SEM(n), where n is the number of independent experiments
T-type channels are characterized by a EEDD ring in the Selectivity Filter (SF) which differs from the EEEE locus found in high voltage activated channels

Summary

Introduction

Voltage-gated sodium and calcium channels (NaV and CaV channels, respectively) are important mediators of many physiological functions, including propagation of electrical signal, cell excitability, gene transcription, secretion and synaptic transmission, plasticity and muscle contraction [1,2] Understanding of their structurefunction relationships, in particular their ability to select between ions of different size and/or charge, are still poorly understood and remain a major task in membrane biophysics. Computational and experimental studies on bacterial voltage-gated sodium channels (which represent models for understanding selectivity in eukaryotic voltage-gated Na+ and Ca2+ channels) are only beginning to provide insight into cation selectivity and conduction Analysis of their recently discovered crystal structures identified glutamates in the SF, which form a high field strength (HFS) binding site favouring Na+ binding with respect to K+ [6]. It is well-established that eukaryotic voltage gated Na+ and Ca2+

Methods

Results

Conclusion