Electrical manipulation of glycan-phosphatidyl inositol-tethered proteins in planar supported bilayers.
Electric fields have been used to manipulate and concentrate glycan-phosphatidyl inositol (GPI)-tethered proteins in planar supported bilayers. Naturally GPI-linked CD48, along with engineered forms of I-Ek and B7-2, in which their transmembrane domains have been genetically replaced with the GPI linkage, were studied. The proteins were labeled with fluorescently tagged antibodies, allowing the electric field-induced behavior to be followed by epifluorescence microscopy. All three protein complexes were observed to migrate toward the cathode with the B7-2 and CD48, each tethered to the membrane by a single GPI linker, moving significantly faster than the I-Ek, which has two GPI linkers. Patterns scratched into the membrane function as barriers to lateral diffusion and were used to isolate the proteins into highly concentrated corrals. All field-induced concentration profiles were completely reversible, indicating that the supported bilayer provides a stable, fluid environment in which GPI-tethered proteins can be manipulated. The ability to electrically control the spatial distribution of membrane-tethered proteins provides new opportunities for the study of biological membranes and the development of membrane-based devices.
Department of Chemistry, Stanford University, California 94305-5080, USA. jtg@leland.stanford.edu
J T
JT
Groves
C
C
Wülfing
S G
SG
Boxer
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry; D051940:B7-2 Antigen; D001483:Base Sequence; D000071178:CD48 Antigen; D016466:CHO Cells; D006224:Cricetinae; D004586:Electrophoresis; D004594:Electrophysiology; D017261:Glycosylphosphatidylinositols; D000949:Histocompatibility Antigens Class II / Q000737:chemistry; D008051:Lipid Bilayers / Q000737:chemistry*; D008560:Membrane Fluidity; D008562:Membrane Glycoproteins / Q000737:chemistry; D008563:Membrane Lipids / Q000737:chemistry; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000737:chemistry*; D014741:Video Recording
D015703:Antigens, CD; D051940:B7-2 Antigen; D000071178:CD48 Antigen; C000604066:Cd48 protein, mouse; C497965:Cd86 protein, mouse; D017261:Glycosylphosphatidylinositols; D000949:Histocompatibility Antigens Class II; C024138:I-E-antigen; D008051:Lipid Bilayers; D008562:Membrane Glycoproteins; D008563:Membrane Lipids; D011134:Polysaccharides