Projection structure of NhaA, a secondary transporter from Escherichia coli, at 4.0 Å resolution

Karen A. Williams*, Ulrike Geldmacher-Kaufer, Etana Padan, Shimon Schuldiner, Werner Kühlbrandt

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

115 Scopus citations

Abstract

Electron cryomicroscopy of frozen-hydrated two-dimensional crystals of NhaA, a Na+/H+ antiporter from Escherichia coli predicted to have 12 transmembrane α-helices, has facilitated the calculation of a projection map of NhaA at 4.0 Å resolution. NhaA was homologously expressed in E.coli with a His6 tag, solubilized in dodecyl maltoside and purified in a single step using Ni2+ affinity chromatography. Two-dimensional crystals were obtained after reconstitution of purified protein with E.coli lipids. The projection map reveals that this secondary transporter has a highly asymmetric structure in projection. NhaA exhibits overall dimensions of ~ 38 x 48 Å with a ringshaped density feature probably corresponding to a bundle of tilted helices, adjacent to an elongated region of density containing several peaks indicative of transmembrane helices. Two crystal forms with p22121 symmetry show tightly packed dimers of NhaA which differ in the interactions between adjacent dimers. This work provides the first direct glimpse into the structure of a secondary transporter.

Original languageEnglish
Pages (from-to)3558-3563
Number of pages6
JournalEMBO Journal
Volume18
Issue number13
DOIs
StatePublished - 1 Jul 1999

Keywords

  • Electron cryomicroscopy
  • Escherichia coli
  • Na/H antiporter
  • Two-dimensional crystallization

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