Uncovering the lipidic basis for the preparation of functional nicotinic acetylcholine receptor detergent complexes for structural studies

Sci Rep. 2016 Sep 19;6:32766. doi: 10.1038/srep32766.

Abstract

This study compares the lipid composition, including individual phospholipid molecular species of solubilized nAChR detergent complexes (nAChR-DCs) with those of the bulk lipids from their source, Torpedo californica (Tc) electric tissue. This lipidomic analysis revealed seventy-seven (77) phospholipid species in the Tc tissue. Analysis of affinity-purified nAChR-DCs prepared with C-12 to C-16 phospholipid analog detergents alkylphosphocholine (FC) and lysofoscholine (LFC) demonstrated that nAChR-DCs prepared with FC12, LFC14, and LFC16 contained >60 phospholipids/nAChR, which was more than twice of those prepared with FC14, FC16, and LFC12. Significantly, all the nAChR-DCs lacked ethanolamine and anionic phospholipids, contained only four cholesterol molecules, and a limited number of phospholipid molecular species per nAChR. Upon incorporation into oocytes, FC12 produce significant functionality, whereas LFC14 and LFC16 nAChR-DCs displayed an increased functionality as compared to the crude Tc membrane. All three nAChR-DCs displayed different degrees of alterations in macroscopic activation and desensitization kinetics.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylcholine / pharmacology
  • Animals
  • Catalysis
  • Chromatography, Affinity
  • Chromatography, High Pressure Liquid
  • Chromatography, Thin Layer
  • Detergents / chemistry*
  • Electric Organ / metabolism
  • Electrodes
  • Hydrolysis
  • Lipids / chemistry*
  • Lipids / isolation & purification
  • Oocytes / drug effects
  • Oocytes / metabolism
  • Patch-Clamp Techniques
  • Receptors, Nicotinic / chemistry*
  • Spectrometry, Mass, Electrospray Ionization
  • Tandem Mass Spectrometry
  • Torpedo
  • Xenopus

Substances

  • Detergents
  • Lipids
  • Receptors, Nicotinic
  • Acetylcholine