Analyzing native membrane protein assembly in nanodiscs by combined non-covalent mass spectrometry and synthetic biology
Abstract
Membrane proteins frequently assemble into higher order homo- or hetero-oligomers within their natural lipid environment. This complex formation can modulate their folding, activity as well as substrate selectivity. Non-disruptive methods avoiding critical steps such as membrane disintegration, transfer into artificial environments or chemical modifications are therefore essential to analyze molecular mechanisms of native membrane protein assemblies. The combination of cell-free synthetic biology, nanodisc-technology and non-covalent mass spectrometry provides excellent synergies for the analysis of membrane protein oligomerization within defined membranes. We exemplify our strategy by oligomeric state characterization of various membrane proteins including ion channels, transporters and membrane integrated enzymes assembling up to hexameric complexes. We further indicate a lipid dependent dimer formation of MraY translocase correlating with the enzymatic activity. The detergent free synthesis of membrane protein/nanodisc samples and the analysis by LILBID mass spectrometry provides a versatile platform for the analysis of membrane proteins in a native environment.
Article and author information
Author details
Funding
Deutsche Forschungsgemeinschaft (Collaborative Research Center (SFB) 807)
- Erik Henrich
- Oliver Peetz
- Christopher Hein
European Strategy Forum on Research Infrastructures (Instruct)
- Frank Bernhard
Deutsche Forschungsgemeinschaft (DO545/11)
- Aisha Laguerre
National Institutes of Health (U54GM087519)
- Beate Hoffmann
Max Planck Research School for Structure and Function of Biological Membranes
- Beate Hoffmann
Cluster of Excellence Frankfurt
- Volker Dötsch
- Nina Morgner
European Research Council (European Union's Seventh Framework Programme (FP7/2007-2013)/ ERC Grant agreement n{degree sign} 337)
- Nina Morgner
P4EU
- Frank Bernhard
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Olga Boudker, Weill Cornell Medical College, United States
Version history
- Received: August 25, 2016
- Accepted: January 4, 2017
- Accepted Manuscript published: January 9, 2017 (version 1)
- Version of Record published: January 26, 2017 (version 2)
Copyright
© 2017, Henrich et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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