Ion Transport and Inhibitor Binding by human NHE1: Insights from Molecular Dynamics Simulations and Free Energy Calculations

30 August 2023, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


The human Na+/H+ exchanger one (hNHE1) plays a crucial role in maintaining intracellular pH by regulating the electroneutral exchange of a single intracellular H+ for one extracellular Na+ across the plasma membrane. Understanding the molecular mechanisms governing ion transport and the binding of inhibitors is of importance in the development of anticancer therapeutics targeting hNHE1. In this context, we performed molecular dynamics (MD) simulations based on the recent cryo-electron microscopy (cryo-EM) structures of outward and inward-facing conformations of hNHE1. These simulations allowed us to explore the dynamics of the protein, examine the ion- translocation pore and confirm that Asp267 is the ion-binding residue. Our free energy calculations suggest that Na+ and K+ bind similarly at the ion-binding site. Consequently, Na+ over K+ selectivity cannot be solely explained by differences in ion binding. Our MD simulations involving hNHE1 inhibitors (cariporide and amiloride analogues), showed maintained stable interactions with Asp267 and Glu346. Our study highlights the importance of the salt bridge between the positively charged acylguanidine moiety and Asp267, which appears to play a role in the competitive inhibitory mechanism for this class of inhibitors. Our computational study provides a detailed mechanistic interpretation of experimental data and serves the basis of future structure-based inhibitor design.


Na+/H+ exchanger 1
MD simulations

Supplementary materials

Supporting Information
Figures S1 and S2 with models of hNHE1, Tables S1 and S2 with simulation details, Tables S3 and S4 and Figures S3 – S9 with detailed results on homology models (supplementary section on homology modelling), Table S5 with structures and IC50 values of inhibitors, Figures S10− S18 and Table S6 with detailed simulation results on cryo-EM structures.


Comments are not moderated before they are posted, but they can be removed by the site moderators if they are found to be in contravention of our Commenting Policy [opens in a new tab] - please read this policy before you post. Comments should be used for scholarly discussion of the content in question. You can find more information about how to use the commenting feature here [opens in a new tab] .
This site is protected by reCAPTCHA and the Google Privacy Policy [opens in a new tab] and Terms of Service [opens in a new tab] apply.