title: Assay of Flippase Activity in Proteoliposomes Using Fluorescent Lipid Derivatives.
abstract: Specific membrane proteins, termed lipid flippases, play a central role in facilitating the movement of lipids across cellular membranes. In this protocol, we describe the reconstitution of ATP-driven lipid flippases in liposomes and the analysis of their in vitro flippase activity based on the use of fluorescent lipid derivatives. Working with purified and reconstituted systems provides a well-defined experimental setup and allows to directly characterize these membrane proteins at the molecular level.
authors.affiliation: Department of Plant and Environmental Sciences, Centre for Membrane Pumps in Cells and Disease (PUMPKIN), University of Copenhagen, Thorvaldsensvej 40, 1871, Frederiksberg C, Denmark.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Marek
authors.affiliation: Department of Plant and Environmental Sciences, Centre for Membrane Pumps in Cells and Disease (PUMPKIN), University of Copenhagen, Thorvaldsensvej 40, 1871, Frederiksberg C, Denmark. tgp@plen.ku.dk.
authors.forename: Thomas
authors.initials: T
authors.lastname: Günther-Pomorski
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D004227:Dithionite / Q000737:chemistry; D005453:Fluorescence; D008055:Lipids / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008967:Molecular Biology / Q000379:methods*; D011510:Proteolipids / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018528:ATP-Binding Cassette Transporters; D008055:Lipids; D008565:Membrane Proteins; D011510:Proteolipids; C020485:proteoliposomes; D004227:Dithionite; D000255:Adenosine Triphosphate
keywords: ABC transporter; Dithionite; Fluorescence; NBD-lipid; P-type ATPase; Reconstitution