Interaction of Human Chloride Intracellular Channel Protein 1 (CLIC1) with Lipid Bilayers: A Fluorescence Study.
Chloride intracellular channel protein 1 (CLIC1) is very unusual as it adopts a soluble glutathione S-transferase-like canonical fold but can also autoinsert into lipid bilayers to form an ion channel. The conversion between these forms involves a large, but reversible, structural rearrangement of the CLIC1 module. The only identified environmental triggers controlling the metamorphic transition of CLIC1 are pH and oxidation. Until now, there have been no high-resolution structural data available for the CLIC1 integral membrane state, and consequently, a limited understanding of how CLIC1 unfolds and refolds across the bilayer to form a membrane protein with ion channel activity exists. Here we show that fluorescence spectroscopy can be used to establish the interaction and position of CLIC1 in a lipid bilayer. Our method employs a fluorescence energy transfer (FRET) approach between CLIC1 and a dansyl-labeled lipid analogue to probe the CLIC1-lipid interface. Under oxidizing conditions, a strong FRET signal between the single tryptophan residue of CLIC1 (Trp35) and the dansyl-lipid analogue was detected. When considering the proportion of CLIC1 interacting with the lipid bilayer, as estimated by fluorescence quenching experiments, the FRET distance between Trp35 and the dansyl moiety on the membrane surface was determined to be ∼15 Å. This FRET-detected interaction provides direct structural evidence that CLIC1 associates with membranes. The results presented support the current model of an oxidation-driven interaction of CLIC1 with lipid bilayers and also propose a membrane anchoring role for Trp35.
Department of Chemistry and Biomolecular Sciences, Macquarie University , Sydney, New South Wales 2109, Australia.
Joanna E
JE
Hare
Astbury Centre for Structural Molecular Biology and School of Molecular and Cellular Biology, University of Leeds , Leeds LS29JT, United Kingdom.
Sophia C
SC
Goodchild
St Vincent's Centre for Applied Medical Research, St Vincent's Hospital , Sydney, New South Wales 2010, Australia.
Samuel N
SN
Breit
School of Physics, University of New South Wales , Sydney, New South Wales 2052, Australia.
Paul M G
PM
Curmi
Department of Chemistry and Biomolecular Sciences, Macquarie University , Sydney, New South Wales 2109, Australia.
Louise J
LJ
Brown
D002462:Cell Membrane / Q000378:metabolism*; D018118:Chloride Channels / Q000737:chemistry* / Q000378:metabolism*; D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D010084:Oxidation-Reduction; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
C501966:CLIC1 protein, human; D018118:Chloride Channels; D008051:Lipid Bilayers