title: Concentrating membrane proteins using asymmetric traps and AC electric fields.
abstract: Membrane proteins are key components of the plasma membrane and are responsible for control of chemical ionic gradients, metabolite and nutrient transfer, and signal transduction between the interior of cells and the external environment. Of the genes in the human genome, 30% code for membrane proteins (Krogh et al. J. Mol. Biol.2001, 305, 567). Furthermore, many FDA-approved drugs target such proteins (Overington et al. Nat. Rev. Drug Discovery 2006, 5, 993). However, the structure-function relationships of these are notably sparse because of difficulties in their purification and handling outside of their membranous environment. Methods that permit the manipulation of membrane components while they are still in the membrane would find widespread application in separation, purification, and eventual structure-function determination of these species (Poo et al. Nature 1977, 265, 602). Here we show that asymmetrically patterned supported lipid bilayers in combination with AC electric fields can lead to efficient manipulation of charged components. We demonstrate the concentration and trapping of such components through the use of a "nested trap" and show that this method is capable of yielding an approximately 30-fold increase in the average protein concentration. Upon removal of the field, the material remains trapped for several hours as a result of topographically restricted diffusion. Our results indicate that this method can be used for concentrating and trapping charged membrane components while they are still within their membranous environment. We anticipate that our approach could find widespread application in the manipulation and study of membrane proteins.
authors.affiliation: School of Physics & Astronomy, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, UK.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Cheetham
authors.forename: Jonathan P
authors.initials: JP
authors.lastname: Bramble
authors.forename: Duncan G G
authors.initials: DG
authors.lastname: McMillan
authors.forename: Lukasz
authors.initials: L
authors.lastname: Krzeminski
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Han
authors.forename: Benjamin R G
authors.initials: BR
authors.lastname: Johnson
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Bushby
authors.forename: Peter D
authors.initials: PD
authors.lastname: Olmsted
authors.forename: Lars J C
authors.initials: LJ
authors.lastname: Jeuken
authors.forename: Sophie J
authors.initials: SJ
authors.lastname: Marritt
authors.forename: Julea N
authors.initials: JN
authors.lastname: Butt
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Evans
mesh_terms: D002462:Cell Membrane / Q000737:chemistry; D004560:Electricity; D008051:Lipid Bilayers / Q000737:chemistry; D008565:Membrane Proteins / Q000302:isolation & purification*
chemical_list: D008051:Lipid Bilayers; D008565:Membrane Proteins