The crossing of two unwound transmembrane regions that is the hallmark of the NhaA structural fold is critical for antiporter activity

Abraham Rimon, Hadar Amartely, Etana Padan*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Cell pH and Na+ homeostasis requires Na+/H+ antiporters. The crystal structure of NhaA, the main Escherichia coli Na+/H+ antiporter, revealed a unique NhaA structural fold shared by prokaryotic and eukaryotic membrane proteins. Out of the 12 NhaA transmembrane segments (TMs), TMs III–V and X–XII are topologically inverted repeats with unwound TMs IV and XI forming the X shape characterizing the NhaA fold. We show that intramolecular cross-linking under oxidizing conditions of a NhaA mutant with two Cys replacements across the crossing (D133C-T340C) inhibits antiporter activity and impairs NhaA-dependent cell growth in high-salts. The affinity purified D133C-T340C protein binds Li+ (the Na+ surrogate substrate of NhaA) under reducing conditions. The cross-linking traps the antiporter in an outward-facing conformation, blocking the antiport cycle. As many secondary transporters are found to share the NhaA fold, including some involved in human diseases, our data have importance for both basic and clinical research.

Original languageEnglish
Article number5915
JournalScientific Reports
Volume14
Issue number1
DOIs
StatePublished - Dec 2024

Bibliographical note

Publisher Copyright:
© The Author(s) 2024.

Keywords

  • Escherichia coli
  • Membrane transport activity
  • NhaA
  • NhaA structural fold
  • Proton antiporter
  • Sodium

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