A rapid method for assessing lipid:protein and detergent:protein ratios in membrane-protein crystallization

Acta Crystallogr D Biol Crystallogr. 2003 Jan;59(Pt 1):77-83. doi: 10.1107/s0907444902019236. Epub 2002 Dec 19.


A simple procedure for rapidly measuring lipid:protein ratios and detergent concentrations at different stages of the solubilization, purification and crystallization of membrane proteins has been developed. Fourier-transform infrared spectra recorded from 10 micro l aliquots of solution using a single-bounce diamond-attenuated total reflectance apparatus exhibit characteristic bands arising from the vibrations of lipid, protein and detergent. Lipid:protein molar ratios as low as 5:1 (for a protein with a molecular weight of 300 kDa) are determined by comparing the ratio of the integrated intensity of the lipid ester carbonyl band near 1740 cm(-1) with the protein amide I band near 1650 cm(-1). Detergent concentrations at levels well below the critical micellar concentration of most detergents are determined by comparing the integrated intensities of the detergent vibrations, particularly in the 1200-1000 cm(-1) region, with a standard curve. Protein amide I band-shape analysis provides insight into the effects of detergents on protein secondary structure. The importance of monitoring detergent concentration changes during simple procedures, such as the concentration of a membrane protein by ultrafiltration, is demonstrated. This analytical tool has been used to rapidly establish protocols for minimizing lipid and detergent levels in solubilized membrane-protein samples.

Publication types

  • Comparative Study

MeSH terms

  • Amides / chemistry
  • Crystallization / methods*
  • Filtration / methods
  • Membrane Proteins / analysis
  • Membrane Proteins / chemistry*
  • Micelles
  • Muramidase / analysis
  • Muramidase / chemistry*
  • Phosphatidylcholines / analysis
  • Phosphatidylcholines / chemistry*
  • Protein Structure, Secondary
  • Solubility
  • Spectroscopy, Fourier Transform Infrared
  • Surface-Active Agents / analysis
  • Surface-Active Agents / chemistry*


  • Amides
  • Membrane Proteins
  • Micelles
  • Phosphatidylcholines
  • Surface-Active Agents
  • Muramidase
  • 1-palmitoyl-2-oleoylphosphatidylcholine