title: Dynamic membrane protein topological switching upon changes in phospholipid environment.
abstract: A fundamental objective in membrane biology is to understand and predict how a protein sequence folds and orients in a lipid bilayer. Establishing the principles governing membrane protein folding is central to understanding the molecular basis for membrane proteins that display multiple topologies, the intrinsic dynamic organization of membrane proteins, and membrane protein conformational disorders resulting in disease. We previously established that lactose permease of Escherichia coli displays a mixture of topological conformations and undergoes postassembly bidirectional changes in orientation within the lipid bilayer triggered by a change in membrane phosphatidylethanolamine content, both in vivo and in vitro. However, the physiological implications and mechanism of dynamic structural reorganization of membrane proteins due to changes in lipid environment are limited by the lack of approaches addressing the kinetic parameters of transmembrane protein flipping. In this study, real-time fluorescence spectroscopy was used to determine the rates of protein flipping in the lipid bilayer in both directions and transbilayer flipping of lipids triggered by a change in proteoliposome lipid composition. Our results provide, for the first time to our knowledge, a dynamic picture of these events and demonstrate that membrane protein topological rearrangements in response to lipid modulations occur rapidly following a threshold change in proteoliposome lipid composition. Protein flipping was not accompanied by extensive lipid-dependent unfolding of transmembrane domains. Establishment of lipid bilayer asymmetry was not required but may accelerate the rate of protein flipping. Membrane protein flipping was found to accelerate the rate of transbilayer flipping of lipids.
authors.affiliation: Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas Health Science Center-Medical School, Houston, TX 77030 heidi.vitrac@uth.tmc.edu william.dowhan@uth.tmc.edu.
authors.forename: Heidi
authors.initials: H
authors.lastname: Vitrac
authors.affiliation: Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas Health Science Center-Medical School, Houston, TX 77030.
authors.forename: David M
authors.initials: DM
authors.lastname: MacLean
authors.affiliation: Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas Health Science Center-Medical School, Houston, TX 77030.
authors.forename: Vasanthi
authors.initials: V
authors.lastname: Jayaraman
authors.affiliation: Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas Health Science Center-Medical School, Houston, TX 77030.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Bogdanov
authors.affiliation: Department of Biochemistry and Molecular Biology and Center for Membrane Biology, University of Texas Health Science Center-Medical School, Houston, TX 77030 heidi.vitrac@uth.tmc.edu william.dowhan@uth.tmc.edu.
authors.forename: William
authors.initials: W
authors.lastname: Dowhan
mesh_terms: D004926:Escherichia coli / Q000201:enzymology; D031541:Fluorescence Resonance Energy Transfer; D008051:Lipid Bilayers; D008565:Membrane Proteins / Q000737:chemistry*; D026901:Membrane Transport Proteins / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry*; D011510:Proteolipids; D013050:Spectrometry, Fluorescence
chemical_list: D008051:Lipid Bilayers; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010743:Phospholipids; D011510:Proteolipids; C020485:proteoliposomes; C021499:lactose permease
keywords: lipid–protein interactions; membrane protein; phospholipids; real-time FRET; topology