Functional Reconstitution of an ABC Transporter in Nanodiscs for Use in Electron Paramagnetic Resonance Spectroscopy
- 25 June 2010
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of the American Chemical Society
- Vol. 132 (28), 9513-9515
- https://doi.org/10.1021/ja104047c
Abstract
Electron paramagnetic resonance (EPR) spectroscopy is a powerful biophysical technique for study of the structural dynamics of membrane proteins. Many of these proteins interact with ligands or proteins on one or both sides of the membrane. Membrane proteins are typically reconstituted in proteoliposomes to observe their function in a physiologically relevant environment. However, membrane proteins can insert into liposomes in two different orientations, and surfaces facing the lumen of the vesicle can be inaccessible to ligands. This heterogeneity can lead to subpopulations that do not respond to ligand binding, complicating EPR spectral analysis, particularly for distance measurements. Using the well-characterized maltose transporter, an ATP binding cassette (ABC) transporter that interacts with ligands on both sides of the membrane, we provide evidence that reconstitution into nanodiscs, which are soluble disk-shaped phospholipid bilayers, is an ideal solution to these problems. We describe the functional reconstitution of the maltose transporter into nanodiscs and demonstrate that this system is ideally suited to study conformational changes and intramolecular distances by EPR.Keywords
This publication has 20 references indexed in Scilit:
- Increased Sensitivity and Extended Range of Distance Measurements in Spin-Labeled Membrane Proteins: Q-Band Double Electron-Electron Resonance and Nanoscale BilayersBiophysical Journal, 2010
- Transmembrane Signaling in the Maltose ABC Transporter MalFGK2-EJournal of Biological Chemistry, 2009
- The nanodisc: a novel tool for membrane protein studiesBiological Chemistry, 2009
- Both maltose-binding protein and ATP are required for nucleotide-binding domain closure in the intact maltose ABC transporterProceedings of the National Academy of Sciences, 2008
- Distance measurements in the borderline region of applicability of CW EPR and DEER: A model study on a homologous series of spin-labelled peptidesJournal of Magnetic Resonance, 2007
- Catalytic Cycle of ATP Hydrolysis by P-Glycoprotein: Evidence for Formation of the E·S Reaction Intermediate with ATP-γ-S, a Nonhydrolyzable Analogue of ATPBiochemistry, 2007
- Crystal structure of a catalytic intermediate of the maltose transporterNature, 2007
- Cooperativity in Cytochrome P450 3A4Journal of Biological Chemistry, 2007
- Applications of Phospholipid Bilayer Nanodiscs in the Study of Membranes and Membrane ProteinsBiochemistry, 2007
- Directed Self-Assembly of Monodisperse Phospholipid Bilayer Nanodiscs with Controlled SizeJournal of the American Chemical Society, 2004