Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 112, Issue 45, Pages 13874-13879Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1512994112
Keywords
membrane protein; topology; lipid-protein interactions; phospholipids; real-time FRET
Categories
Funding
- NIH [R37 GM 20478, R01 GM 113212, R01 GM 094246-04]
- John Dunn Research Foundation
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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 formembrane 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.
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