A facile, sensitive and quantitative membrane-binding assay for proteins

Traffic. 2020 Mar;21(3):297-305. doi: 10.1111/tra.12719. Epub 2019 Dec 25.

Abstract

Soluble proteins that bind membranes function in numerous cellular pathways yet facile, sensitive and quantitative methods that complement and improve sensitivity of widely used liposomes-based assays remain unavailable. Here, we describe the utility of a photoactivable fluorescent lipid as a generic reporter of protein-membrane interactions. When incorporated into liposomes and exposed to ultraviolet (UV), proteins bound to liposomes become crosslinked with the fluorescent lipid and can be readily detected and quantitated by in-gel fluorescence analysis. This modification obviates the requirement for high-speed centrifugation spins common to most liposome-binding assays. We refer to this assay as Proximity-based Labeling of Membrane-Associated Proteins (PLiMAP).

Keywords: BODIPY; diazirine; fluorescence; in-gel analysis; liposomes; phosphoinositides; photoactivation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Lipids*
  • Liposomes*
  • Membrane Proteins*
  • Membranes

Substances

  • Lipids
  • Liposomes
  • Membrane Proteins