Probing the ATP hydrolysis cycle of the ABC multidrug transporter LmrA by pulsed EPR spectroscopy

J Am Chem Soc. 2012 Apr 4;134(13):5857-62. doi: 10.1021/ja211007t. Epub 2012 Mar 21.


Members of the ATP binding cassette (ABC) transporter superfamily translocate various types of molecules across the membrane at the expense of ATP. This requires cycling through a number of catalytic states. Here, we report conformational changes throughout the catalytic cycle of LmrA, a homodimeric multidrug ABC transporter from L. lactis. Using site-directed spin labeling and pulsed electron-electron double resonance (PELDOR/DEER) spectroscopy, we have probed the reorientation of the nucleotide binding domains and transmembrane helix 6 which is of particular relevance to drug binding and part of the dimerization interface. Our data show that LmrA samples a very large conformational space in its apo state, which is significantly reduced upon nucleotide binding. ATP binding but not hydrolysis is required to trigger this conformational change, which results in a relatively fixed orientation of both the nucleotide binding domains and transmembrane helices 6. This orientation is maintained throughout the ATP hydrolysis cycle until the protein cycles back to its apo state. Our data present strong evidence that switching between two dynamically and structurally distinct states is required for substrate translocation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism*
  • Apoproteins / chemistry
  • Apoproteins / genetics
  • Apoproteins / metabolism
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Biocatalysis
  • Electron Spin Resonance Spectroscopy
  • Hydrolysis
  • Lactococcus lactis
  • Models, Molecular
  • Multidrug Resistance-Associated Proteins / chemistry*
  • Multidrug Resistance-Associated Proteins / genetics
  • Multidrug Resistance-Associated Proteins / metabolism*
  • Mutation
  • Protein Structure, Secondary


  • Apoproteins
  • Bacterial Proteins
  • LmrA protein, Lactococcus lactis
  • Multidrug Resistance-Associated Proteins
  • Adenosine Triphosphate