Using a SMALP platform to determine a sub-nm single particle cryo-EM membrane protein structure

• Published in: Biochimica et biophysica acta Vol. 1860; no. 2; pp. 378 - 383
• Main Authors: Parmar, Mayuriben, Rawson, Shaun, Scarff, Charlotte A., Goldman, Adrian, Dafforn, Timothy R., Muench, Stephen P., Postis, Vincent L.G.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.02.2018; Elsevier Pub. Co
• Subjects: AcrB; Cryoelectron Microscopy - methods; Crystallography, X-Ray; Electron microscopy; Escherichia coli - metabolism; Escherichia coli - ultrastructure; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - ultrastructure; Lipid Bilayers - chemistry; Maleates - chemistry; Membrane proteins; Membrane Proteins - chemistry; Membrane Proteins - ultrastructure; Models, Molecular; Multidrug Resistance-Associated Proteins - chemistry; Multidrug Resistance-Associated Proteins - ultrastructure; Polystyrenes - chemistry; Protein Conformation; Protein Multimerization; Proteolipids - chemistry; Proteolipids - ultrastructure; SMALP; Electron microscopy; AcrB; SMALP; Membrane proteins
• ISSN: 0005-2736; 0006-3002; 1879-2642; 1878-2434
• DOI: 10.1016/j.bbamem.2017.10.005

The field of membrane protein structural biology has been revolutionized over the last few years with a number of high profile structures being solved using cryo-EM including Piezo, Ryanodine receptor, TRPV1 and the Glutamate receptor. Further developments in the EM field hold the promise of even greater progress in terms of greater resolution, which for membrane proteins is still typically within the 4–7Å range. One advantage of a cryo-EM approach is the ability to study membrane proteins in more “native” like environments for example proteoliposomes, amphipols and nanodiscs. Recently, styrene maleic acid co-polymers (SMA) have been used to extract membrane proteins surrounded by native lipids (SMALPs) maintaining a more natural environment. We report here the structure of the Escherichia coli multidrug efflux transporter AcrB in a SMALP scaffold to sub-nm resolution, with the resulting map being consistent with high resolution crystal structures and other EM derived maps. However, both the C-terminal helix (TM12) and TM7 are poorly defined in the map. These helices are at the exterior of the helical bundle and form the greater interaction with the native lipids and SMA polymer and may represent a more dynamic region of the protein. This work shows the promise of using an SMA approach for single particle cryo-EM studies to provide sub-nm structures. [Display omitted] •The first sub nm single particle reconstruction of a membrane protein using a SMALP scaffold•The ability to maintain a native like environment for structural studies holds great promise.•The resulting map is consistent with high resolution crystal structures and other EM derived maps.