Fluorescence quenching methods to study lipid-protein interactions.
This unit describes how fluorescence quenching methods can be used to determine binding constants for phospholipids binding to intrinsic membrane proteins. Reconstitution of a Trp-containing intrinsic membrane protein with bromine-containing phospholipids leads to quenching of the Trp fluorescence of the protein; the extent of quenching depends on the strength of binding of the phospholipid to the protein. Protocols are included for the synthesis of bromine-containing phospholipids from phospholipids containing carbon-carbon double bonds in their fatty acyl chains and for the reconstitution of membrane proteins into bilayers containing bromine-containing phospholipids. Details are included on data analysis, including equations and software that can be used for fitting the fluorescence quenching data.
Carney
Joanne
J
University of Southampton, Southampton, United Kingdom.
East
J Malcolm
JM
Mall
Sanjay
S
Marius
Phedra
P
Powl
Andrew M
AM
Wright
J Neville
JN
Lee
Anthony G
AG
D001966:Bromine / Q000737:chemistry; D005453:Fluorescence; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D010743:Phospholipids / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding
D008565:Membrane Proteins; D010743:Phospholipids; D001966:Bromine