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Biochim Biophys Acta Biomembr. 2018 Feb;1860(2):378-383. doi: 10.1016/j.bbamem.2017.10.005. Epub 2017 Oct 6.

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

Author information

1
Biomedicine Research Group, Faculty of Health and Social Sciences, Leeds Beckett University, LS1 3HE, UK; School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, LS2 9JT, UK.
2
School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, LS2 9JT, UK; Astbury Centre for Structural Molecular Biology, University of Leeds, LS2 9JT, UK.
3
School of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, UK; Astbury Centre for Structural Molecular Biology, University of Leeds, LS2 9JT, UK.
4
School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, LS2 9JT, UK; Astbury Centre for Structural Molecular Biology, University of Leeds, LS2 9JT, UK; Department of Biosciences, Division of Biochemistry, University of Helsinki, Helsinki, Finland.
5
School of Biosciences, University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK.
6
School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, LS2 9JT, UK; Astbury Centre for Structural Molecular Biology, University of Leeds, LS2 9JT, UK. Electronic address: s.p.muench@leeds.ac.uk.
7
Biomedicine Research Group, Faculty of Health and Social Sciences, Leeds Beckett University, LS1 3HE, UK; School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, LS2 9JT, UK. Electronic address: V.L.Postis@leedsbeckett.ac.uk.

Abstract

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.

KEYWORDS:

AcrB; Electron microscopy; Membrane proteins; SMALP

PMID:
28993151
PMCID:
PMC5780298
DOI:
10.1016/j.bbamem.2017.10.005
[Indexed for MEDLINE]
Free PMC Article

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