Slow Phospholipid Exchange between a Detergent-Solubilized Membrane Protein and Lipid-Detergent Mixed Micelles: Brominated Phospholipids as Tools to Follow Its Kinetics

PLoS One. 2017 Jan 24;12(1):e0170481. doi: 10.1371/journal.pone.0170481. eCollection 2017.


Membrane proteins are largely dependent for their function on the phospholipids present in their immediate environment, and when they are solubilized by detergent for further study, residual phospholipids are critical, too. Here, brominated phosphatidylcholine, a phospholipid which behaves as an unsaturated phosphatidylcholine, was used to reveal the kinetics of phospholipid exchange or transfer from detergent mixed micelles to the environment of a detergent-solubilized membrane protein, the paradigmatic P-type ATPase SERCA1a, in which Trp residues can experience fluorescence quenching by bromine atoms present on phospholipid alkyl chains in their immediate environment. Using dodecylmaltoside as the detergent, exchange of (brominated) phospholipid was found to be much slower than exchange of detergent under the same conditions, and also much slower than membrane solubilization, the latter being evidenced by light scattering changes. The kinetics of this exchange was strongly dependent on temperature. It was also dependent on the total concentration of the mixed micelles, revealing the major role for such exchange of the collision of detergent micelles with the detergent-solubilized protein. Back-transfer of the brominated phospholipid from the solubilized protein to the detergent micelle was much faster if lipid-free DDM micelles instead of mixed micelles were added for triggering dissociation of brominated phosphatidylcholine from the solubilized protein, or in the additional presence of C12E8 detergent during exchange, also emphasizing the role of the chemical nature of the micelle/protein interface. This protocol using brominated lipids appears to be valuable for revealing the possibly slow kinetics of phospholipid transfer to or from detergent-solubilized membrane proteins. Independently, continuous recording of the activity of the protein can also be used in some cases to correlate changes in activity with the exchange of a specific phospholipid, as shown here by using the Drs2p/Cdc50p complex, a lipid flippase with specific binding sites for lipids.

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Animals
  • Detergents / pharmacology*
  • Diffusion
  • Fluorometry
  • Glucosides / pharmacology
  • Halogenation
  • Kinetics
  • Membrane Lipids / metabolism*
  • Membrane Proteins / drug effects
  • Membrane Proteins / metabolism*
  • Micelles*
  • Phosphatidylcholines / metabolism*
  • Phospholipids / metabolism*
  • Rabbits
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / drug effects
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / metabolism
  • Solubility
  • Temperature


  • Detergents
  • Glucosides
  • Membrane Lipids
  • Membrane Proteins
  • Micelles
  • Phosphatidylcholines
  • Phospholipids
  • dodecyl maltoside
  • Adenosine Triphosphate
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases

Grants and funding

This work was funded by a grant from the Agence Nationale de la Recherche (ANR, to GL ('AsymLip', ANR-14-CE09-0022) and by the French Infrastructure for Integrated Structural Biology (FRISBI,, ANR-10-INSB-05-01). This work was also supported by the Centre National de la Recherche Scientifique (CNRS). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.