Monomers of the NhaA Na+/H+ antiporter of Escherichia coli are fully functional yet dimers are beneficial under extreme stress conditions at alkaline pH in the presence of Na+ or Li+

Abraham Rimon, Tzvi Tzubery, Etana Padan*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

63 Scopus citations

Abstract

NhaA, the Na+/H+ antiporter of Escherichia coli, exists in the native membrane as a homodimer of which two monomers have been suggested to be attached by a β-hairpin at the periplasmic side of the membrane. Constructing a mutant deleted of the β-hairpin, NhaA/Δ(Pro45-Asn58), revealed that in contrast to the dimeric mobility of native NhaA, the mutant has the mobility of a monomer in a blue native gel. Intermolecular cross-linking that monitors dimers showed that the mutant exists only as monomers in the native membrane, proteoliposomes, and when purified in β-dodecyl maltoside micelles. Furthermore, pull-down experiments revealed that, whereas as expected for a dimer, hemagglutinin-tagged wild-type NhaA co-purified with His-tagged NhaA on a Ni2+-NTA affinity column, a similar version of the mutant did not. Remarkably, under routine stress conditions (0.1 M LiCl, pH 7 or 0.6 M NaCl, pH 8.3), the monomeric form of NhaA is fully functional. It conferred salt resistance to NhaA- and NhaB-deleted cells, and whether in isolated membrane vesicles or reconstituted into proteoliposomes exhibited Na+/H+ antiporter activity and pH regulation very similar to wild-type dimers. Remarkably, under extreme stress conditions (0.1 M LiCl or 0.7 M NaCl at pH 8.5), the dimeric native NhaA was much more efficient than the monomeric mutant in conferring extreme stress resistance.

Original languageEnglish
Pages (from-to)26810-26821
Number of pages12
JournalJournal of Biological Chemistry
Volume282
Issue number37
DOIs
StatePublished - 14 Sep 2007

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