Article
- The EMBO Journal (2008) 27, 17 - 26
- doi:10.1038/sj.emboj.7601951
Published online: 6 December 2007
Subject Categories:
Parallel topology of genetically fused EmrE homodimers
Sonia Steiner-Mordoch1,2, Misha Soskine1,2, Dalia Solomon1, Dvir Rotem1,3, Ayala Gold1, Michal Yechieli1, Yoav Adam1 and Shimon Schuldiner1
- Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
Correspondence to:
Shimon Schuldiner, Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Silberman Bldg 3-345, Jerusalem, Givat Ram 91904, Israel. Tel.: +972 2 658 5992; Fax: +972 2 563 4625; E-mail: Shimon.Schuldiner@huji.ac.il
2These authors contributed equally to this work
3Present address: Department of Chemistry, Oxford University, Oxford, UK
Received 24 July 2007; Accepted 14 November 2007
Abstract
EmrE is a small H+-coupled multidrug transporter in Escherichia coli. Claims have been made for an antiparallel topology of this homodimeric protein. However, our own biochemical studies performed with detergent-solubilized purified protein support a parallel topology of the protomers. We developed an alternative approach to constrain the relative topology of the protomers within the dimer so that their activity can be assayed also in vivo before biochemical handling. Tandem EmrE was built with two identical monomers genetically fused tail to head (C-terminus of the first to N-terminus of the second monomer) with hydrophilic linkers of varying length. All the constructs conferred resistance to ethidium by actively removing it from the cytoplasm. The purified proteins bound substrate and transported methyl viologen into proteoliposomes by a proton-dependent mechanism. A tandem where one of the essential glutamates was replaced with glutamine transported only monovalent substrates and displayed a modified stoichiometry. The results support a parallel topology of the protomers in the functional dimer. The implications regarding insertion and evolution of membrane proteins are discussed.
Keywords:
- ion-coupled transport,
- membrane protein evolution,
- membrane protein structure,
- membrane protein topology,
- multidrug transporters
MORE ARTICLES LIKE THIS
These links to content published by NPG are automatically generated
REVIEWS
Multiple molecular mechanisms for multidrug resistance transporters
Nature Review (12 Apr 2007)
NEWS AND VIEWS
Flip-flopping membrane proteins
Nature Structural & Molecular Biology News and Views (01 Feb 2006)
RESEARCH
The EMBO Journal Article (01 Dec 2003)
The EMBO Journal Article (01 Dec 2003)
A membrane-embedded glutamate is required for ligand binding to the multidrug transporter EmrE
The EMBO Journal Article (17 Jan 2000)
