CLIC1 inserts from the aqueous phase into phospholipid membranes, where it functions as an anion channel.
CLIC1 is a member of the CLIC family of proteins, which has been shown to demonstrate chloride channel activity when reconstituted in phospholipid vesicles. CLIC1 exists in cells as an integral membrane protein and as a soluble cytoplasmic protein, implying that CLIC1 might cycle between membrane-inserted and soluble forms. CLIC1 was purified and detergent was removed, yielding an aqueous solution of essentially pure protein. Pure CLIC1 was mixed with vesicles, and chloride permeability was assessed with a chloride efflux assay and with planar lipid bilayer techniques. Soluble CLIC1 confers anion channel activity to preformed membranes that is indistinguishable from the previously reported activity resulting from reconstitution of CLIC1 into membranes by detergent dialysis. The activity is dependent on the amount of CLIC1 added, appears rapidly on mixing of protein and lipid, is inhibited by indanyloxyacetic acid-94, N-ethylmaleimide, and glutathione, is inactivated by heat, and shows sensitivity to pH and to membrane lipid composition. We conclude that CLIC1 in the absence of detergent spontaneously inserts into preformed membranes, where it can function as an anion-selective channel.
Tulk
Barry M
BM
Department of Medicine, St. Louis University and St. Louis Veterans Affairs Medical Center, St. Louis, Missouri 63106, USA.
Kapadia
Shefalee
S
Edwards
John C
JC
D001692:Biological Transport / Q000502:physiology; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004594:Electrophysiology; D006863:Hydrogen-Ion Concentration; D010743:Phospholipids / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D018118:Chloride Channels; D002712:Chlorides; D010743:Phospholipids; D011993:Recombinant Fusion Proteins; D017868:Cyclic AMP-Dependent Protein Kinases