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Scientific article
English

Generation of proteoliposomes from subcellular fractions

Published inElectrophoresis, vol. 18, no. 14, p. 2620-2628
Publication date1997
Abstract

Intracellular membranes are highly dynamic, yet they retain their identity and functional characteristics. Integral membrane proteins, which must confer this specific membrane identity, remain poorly characterized at the biochemical level, largely because detergent-mediated solubilization is required for purification and analysis, and several properties of integral membrane proteins can only be investigated when the molecule is properly embedded in a lipid bilayer. We present a method for the efficient reconstitution into proteoliposomes of integral membrane proteins from subcellular fractions. Integral membrane proteins were identified on high-resolution two-dimensional gels after selective extraction of soluble and peripheral membrane proteins; they accounted for 8% of the number of resolved polypeptides. A reconstitution procedure based on membrane solubilization with dodecyl-octaoxyethylene (C12E8) and subsequent detergent removal with BioBeads SM-2 resulted in the efficient reconstitution of several membrane proteins into proteoliposomes of uniform density. The generated proteoliposomes strongly resemble the starting membrane fraction in protein composition. This reconstitution allows the functional characterization of integral membrane proteins after enrichment and/or specific (immuno)depletion.

eng
Keywords
  • Membrane solubilization
  • Membrane reconstitution
  • Proteoliposome
  • Detergent
  • Integral membrane protein
  • Two-dimensional polyacrylamide gel electrophoresis
  • Endosome
  • Golgi
Citation (ISO format)
ROJO ORTIZ DE LANZAGORTA, Manuel et al. Generation of proteoliposomes from subcellular fractions. In: Electrophoresis, 1997, vol. 18, n° 14, p. 2620–2628. doi: 10.1002/elps.1150181418
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ISSN of the journal0173-0835
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