Molecular Orientation Determination in Nanodiscs at the Single-Molecule Level

Tyler Camp, Kritika Mehta, Stephen G. Sligar, Kai Zhang

Research output: Contribution to journalArticle

Abstract

The function of membrane-bound proteins often depends on their interactions with the lipid bilayer. Bulk absorption-based linear dichroism has been historically used to investigate molecular orientations in the phospholipid bilayer but cannot resolve the actual distribution of molecules embedded in the membrane and is often limited by a poor signal-to-noise ratio. Here, we present single-molecule orientation determination by fluorescence-detected linear dichroism visualization in Nanodisc grids or SOLVING, to determine the molecular orientation of molecules assembled into nanoscale lipid bilayers. We provide a proof-of-concept by using SOLVING to quantitate the orientation distribution of two commonly used fluorescent dyes, DiO and BODIPY, in 10 nm Nanodiscs. Besides confirming the mean orientation determined by bulk absorption measurement, SOLVING provides the actual distribution of orientations and promises to provide key molecular insights into the topology and interactions of multiprotein complexes, such as those observed in intracellular signal transduction.

Original languageEnglish (US)
Pages (from-to)2229-2236
Number of pages8
JournalAnalytical chemistry
Volume92
Issue number2
DOIs
StatePublished - Jan 21 2020

ASJC Scopus subject areas

  • Analytical Chemistry

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