Structure and mechanism of the Mrp complex, an ancient cation/proton antiporter.

Julia Steiner,Leonid Sazanov

doi:10.7554/elife.59407

Abstract

Multiple resistance and pH adaptation (Mrp) antiporters are multi-subunit Na+ (or K+)/H+ exchangers representing an ancestor of many essential redox-driven proton pumps, such as respiratory complex I. The mechanism of coupling between ion or electron transfer and proton translocation in this large protein family is unknown. Here, we present the structure of the Mrp complex from Anoxybacillus flavithermus solved by cryo-EM at 3.0 Å resolution. It is a dimer of seven-subunit protomers with 50 trans-membrane helices each. Surface charge distribution within each monomer is remarkably asymmetric, revealing probable proton and sodium translocation pathways. On the basis of the structure we propose a mechanism where the coupling between sodium and proton translocation is facilitated by a series of electrostatic interactions between a cation and key charged residues. This mechanism is likely to be applicable to the entire family of redox proton pumps, where electron transfer to substrates replaces cation movements.

Highlights

The Na+/H+ antiporters are widely distributed secondary active transporters that use the proton motive force to efflux intracellular sodium ions (Ito et al, 2017)
The His-tagged Multiple resistance and pH adaptation (Mrp) complex from A. flavithermus was recombinantly expressed in the antiporter-deficient E. coli strain KNabc (Goldberg et al, 1987)
A striking electrostatic imbalance of H+ and Na+ modules of Mrp (Figure 2b,c), the arrangement of key residues in the H+ and Na+ pathways, the separation of H+ cross-pathways from the LysTMH7/GluTMH5 coupling points and a remarkable similarity of the MrpAN/MrpD and MrpD/ Na+ coupling points (Figure 4a,b) all collectively suggest that electrostatic interactions are the main driving force in the antiport mechanism

Summary

Introduction

The Na+/H+ antiporters are widely distributed secondary active transporters that use the proton motive force to efflux intracellular sodium ions (Ito et al, 2017). The largest two Mrp subunits, MrpA and MrpD, are homologous to each other, have 14 conserved trans-membrane (TM) helices, and are thought to participate in proton translocation (Mathiesen and Hagerhall, 2003) Their homologues (called antiporter-like subunits, which we will abbreviate to APLS) are found in many proton-pumping protein complexes where they are present in one to three (and recently discovered four [Chadwick et al, 2018]) copies per complex, depending on the energy availability and needs of the organism (Efremov and Sazanov, 2012).

Results

Discussion

Materials and methods