Measuring Membrane Protein Dimerization Equilibrium in Lipid Bilayers by Single-Molecule Fluorescence Microscopy

R. Chadda, J. L. Robertson

Research output: Chapter in Book/Report/Conference proceedingChapter

3 Scopus citations

Abstract

Dimerization of membrane protein interfaces occurs during membrane protein folding and cell receptor signaling. Here, we summarize a method that allows for measurement of equilibrium dimerization reactions of membrane proteins in lipid bilayers, by measuring the Poisson distribution of subunit capture into liposomes by single-molecule photobleaching analysis. This strategy is grounded in the fact that given a comparable labeling efficiency, monomeric or dimeric forms of a membrane protein will give rise to distinctly different photobleaching probability distributions. These methods have been used to verify the dimer stoichiometry of the Fluc F ion channel and the dimerization equilibrium constant of the ClC-ec1 Cl/H+ antiporter in lipid bilayers. This approach can be applied to any membrane protein system provided it can be purified, fluorescently labeled in a quantitative manner, and verified to be correctly folded by functional assays, even if the structure is not yet known.

Original languageEnglish
Title of host publicationMethods in Enzymology
PublisherAcademic Press Inc.
Pages53-82
Number of pages30
DOIs
StatePublished - Jan 1 2016
Externally publishedYes

Publication series

NameMethods in Enzymology
Volume581
ISSN (Print)0076-6879
ISSN (Electronic)1557-7988

Keywords

  • Dimerization
  • Equilibrium
  • Lipid bilayers
  • Membrane protein
  • Photobleaching
  • Poisson
  • Single-molecule microscopy

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