Home LiteratureArticle Details
PMID: 17704177 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

High-resolution structure of a Na+/H+ antiporter dimer obtained by pulsed electron paramagnetic resonance distance measurements.

Biophysical journal ·Vol. 93 ·No. 10 ·2007-11-15 ·Pages 3675-83

Hilger D, Polyhach Y, Padan E, Jung H, Jeschke G

Abstract

Transient or partial formation of complexes between biomacromolecules is a general mechanism used to control cellular functions. Several of these complexes escape structure determination by crystallographic means. We developed a new approach for determining the structure of protein dimers in the native environment (e.g., in the membrane) with high resolution in cases where the structure of the two monomers is known. The approach is based on measurements of distance distributions between spin labels in the range between 2 and 6 nanometers by a pulsed electron paramagnetic resonance technique and explicit modeling of spin label conformations. By applying this method to the membrane protein homodimer of the Na(+)/H(+) antiporter NhaA of Escherichia coli, the structure of the presumably physiological dimer was determined. It reveals two points of contact between the two monomers, with one of them confirming results of earlier cross-linking experiments.

MeSH Terms
Alleles Binding Sites Cryoelectron Microscopy Cysteine/chemistry Databases, Protein Dimerization Electron Spin Resonance Spectroscopy Escherichia coli/metabolism Escherichia coli Proteins/metabolism Hydrogen-Ion Concentration Models, Statistical Molecular Conformation Mutagenesis, Site-Directed Protein Conformation Sodium-Hydrogen Exchangers/chemistry,metabolism
Chemicals
Escherichia coli Proteins NhaA protein, E coli Sodium-Hydrogen Exchangers Cysteine
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Hilger D
Ludwig-Maximilians-Universität München, Department Biologie I, D-80638 Munich, Germany.
Polyhach Y
Padan E
Jung H
Jeschke G
References (30)
30 references, click to expand
  1. Automated analysis of interatomic contacts in proteins.
    Bioinformatics. 1999 Apr;15(4):327-32 PMID: 10320401
  2. Projection structure of NhaA, a secondary transporter from Escherichia coli, at 4.0 A resolution.
    EMBO J. 1999 Jul 1;18(13):3558-63 PMID: 10393172
  3. A pH-dependent conformational change of NhaA Na(+)/H(+) antiporter of Escherichia coli involves loop VIII-IX, plays a role in the pH response of the protein, and is maintained by the pure protein in dodecyl maltoside.
    J Biol Chem. 1999 Aug 27;274(35):24617-24 PMID: 10455127
  4. Three-dimensional structure of the ion-coupled transport protein NhaA.
    Nature. 2000 Jan 6;403(6765):112-5 PMID: 10638764
  5. Dead-time free measurement of dipole-dipole interactions between electron spins.
    J Magn Reson. 2000 Feb;142(2):331-40 PMID: 10648151
  6. Oligomerization of NhaA, the Na+/H+ antiporter of Escherichia coli in the membrane and its functional and structural consequences.
    Biochemistry. 2001 Mar 20;40(11):3403-12 PMID: 11258962
  7. Calculation of rigid-body conformational changes using restraint-driven Cartesian transformations.
    Biophys J. 2001 Nov;81(5):2530-46 PMID: 11606268
  8. Direct conversion of EPR dipolar time evolution data to distance distributions.
    J Magn Reson. 2002 Mar;155(1):72-82 PMID: 11945035
  9. Proximity of cytoplasmic and periplasmic loops in NhaA Na+/H+ antiporter of Escherichia coli as determined by site-directed thiol cross-linking.
    Biochemistry. 2002 Dec 17;41(50):14897-905 PMID: 12475238
  10. HADDOCK: a protein-protein docking approach based on biochemical or biophysical information.
    J Am Chem Soc. 2003 Feb 19;125(7):1731-7 PMID: 12580598
  11. Sensitivity enhancement in pulse EPR distance measurements.
    J Magn Reson. 2004 Jul;169(1):1-12 PMID: 15183350
  12. NhaA of Escherichia coli, as a model of a pH-regulated Na+/H+antiporter.
    Biochim Biophys Acta. 2004 Jul 23;1658(1-2):2-13 PMID: 15282168
  13. The determination of pair distance distributions by pulsed ESR using Tikhonov regularization.
    J Magn Reson. 2005 Feb;172(2):279-95 PMID: 15649755
  14. Assessing oligomerization of membrane proteins by four-pulse DEER: pH-dependent dimerization of NhaA Na+/H+ antiporter of E. coli.
    Biophys J. 2005 Aug;89(2):1328-38 PMID: 15894644
  15. Explicit treatment of spin labels in modeling of distance constraints from dipolar EPR and DEER.
    J Am Chem Soc. 2005 Jul 6;127(26):9334-5 PMID: 15984837
  16. Structure of a Na+/H+ antiporter and insights into mechanism of action and regulation by pH.
    Nature. 2005 Jun 30;435(7046):1197-202 PMID: 15988517
  17. Overproduction and purification of a functional Na+/H+ antiporter coded by nhaA (ant) from Escherichia coli.
    J Biol Chem. 1991 Jun 15;266(17):11289-94 PMID: 1645730
  18. Multiconformation continuum electrostatics analysis of the NhaA Na+/H+ antiporter of Escherichia coli with functional implications.
    Proc Natl Acad Sci U S A. 2006 Feb 21;103(8):2629-34 PMID: 16477015
  19. Distance between a native cofactor and a spin label in the reaction centre of Rhodobacter sphaeroides by a two-frequency pulsed electron paramagnetic resonance method and molecular dynamics simulations.
    J Magn Reson. 2006 Jun;180(2):178-85 PMID: 16515869
  20. Reconstruction of the chemotaxis receptor-kinase assembly.
    Nat Struct Mol Biol. 2006 May;13(5):400-7 PMID: 16622408
  21. Inter-helix distances in lysophospholipid micelle-bound alpha-synuclein from pulsed ESR measurements.
    J Am Chem Soc. 2006 Aug 9;128(31):10004-5 PMID: 16881616
  22. Crucial steps in the structure determination of the Na+/H+ antiporter NhaA in its native conformation.
    J Mol Biol. 2006 Sep 15;362(2):192-202 PMID: 16919297
  23. Distance measurements on spin-labelled biomacromolecules by pulsed electron paramagnetic resonance.
    Phys Chem Chem Phys. 2007 Apr 28;9(16):1895-910 PMID: 17431518
  24. Transmembrane protein structure: spin labeling of bacteriorhodopsin mutants.
    Science. 1990 Jun 1;248(4959):1088-92 PMID: 2160734
  25. The OPLS [optimized potentials for liquid simulations] potential functions for proteins, energy minimizations for crystals of cyclic peptides and crambin.
    J Am Chem Soc. 1988 Mar 1;110(6):1657-66 PMID: 27557051
  26. Determination of the distance between two spin labels attached to a macromolecule.
    Proc Natl Acad Sci U S A. 1995 Aug 29;92(18):8239-43 PMID: 7667275
  27. Calculation of electron paramagnetic resonance spectra from Brownian dynamics trajectories: application to nitroxide side chains in proteins.
    Biophys J. 1996 Oct;71(4):2201-12 PMID: 8889196
  28. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modeling.
    Electrophoresis. 1997 Dec;18(15):2714-23 PMID: 9504803
  29. Unidirectional reconstitution and characterization of purified Na+/proline transporter of Escherichia coli.
    Biochemistry. 1998 Aug 4;37(31):11083-8 PMID: 9693004
  30. A point mutation (G338S) and its suppressor mutations affect both the pH response of the NhaA-Na+/H+ antiporter as well as the growth phenotype of Escherichia coli.
    J Biol Chem. 1998 Oct 9;273(41):26470-6 PMID: 9756882
Article Info
Journal
Biophysical journal
Abbr.
Biophys J
ISSN
0006-3495
DOI
10.1529/biophysj.107.109769
Published
2007-11-15
Epub
2007-00-17
Pages
3675-83
Language
English
Region
United States
NLM ID
0370626
PMCID
PMC2072073
Subset
IM
External Links
PubMed source DOI: 10.1529/biophysj.107.109769 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2