E. coli multidrug transporter MdfA is a monomer

Biochemistry. 2007 May 1;46(17):5200-8. doi: 10.1021/bi602405w. Epub 2007 Apr 4.

Abstract

MdfA is a 410-residue-long secondary multidrug transporter from E. coli. Cells expressing MdfA from a multicopy plasmid exhibit resistance against a diverse group of toxic compounds, including neutral and cationic ones, because of active multidrug export. As a prerequisite for high-resolution structural studies and a better understanding of the mechanism of substrate recognition and translocation by MdfA, we investigated its biochemical properties and overall structural characteristics. To this end, we purified the beta-dodecyl maltopyranoside (DDM)-solubilized protein using a 6-His tag and metal affinity chromatography, and size exclusion chromatography (SE-HPLC). Purified MdfA was analyzed for its DDM and phospholipid (PL) content, and tetraphenylphosphonium (TPP+)-binding activity. The results are consistent with MdfA being an active monomer in DDM solution. Furthermore, an investigation of two-dimensional crystals by electron crystallography and 3D reconstruction lent support to the notion that MdfA may also be monomeric in reconstituted proteoliposomes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, Affinity
  • Chromatography, Gel
  • Chromatography, High Pressure Liquid
  • Crystallization
  • Detergents / chemistry
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / chemistry*
  • Escherichia coli Proteins / chemistry*
  • Membrane Transport Proteins / chemistry*
  • Microscopy, Electron

Substances

  • Detergents
  • Escherichia coli Proteins
  • Mdfa protein, E coli
  • Membrane Transport Proteins