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Expression, purification and crystallisation of the adenosine A2A receptor bound to an engineered Mini G protein
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Carpenter, Byron and Tate, Christopher G. (2017) Expression, purification and crystallisation of the adenosine A2A receptor bound to an engineered Mini G protein. Bio-protocol, 7 (8). e2234. ISSN 2331-8325.
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Official URL: https://doi.org/10.21769/BioProtoc.2234
Abstract
G protein-coupled receptors (GPCRs) promote cytoplasmic signalling by activating heterotrimeric G proteins in response to extracellular stimuli such as light, hormones and nucleosides. Structure determination of GPCR–G protein complexes is central to understanding the precise mechanism of signal transduction. However, these complexes are challenging targets for structural studies due to their conformationally dynamic and inherently transient nature. We recently developed an engineered G protein, mini-Gs, which addressed these problems and allowed the formation of a stable GPCR–G protein complex. Mini-Gs facilitated the structure determination of the human adenosine A2A receptor (A2AR) in its G protein-bound conformation at 3.4 Å resolution. Here, we describe a step by step protocol for the expression and purification of A2AR, and crystallisation of the A2AR–mini-Gs complex.
Item Type: | Journal Article | ||||||||||||
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Subjects: | Q Science > QD Chemistry | ||||||||||||
Divisions: | Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) | ||||||||||||
Journal or Publication Title: | Bio-protocol | ||||||||||||
Publisher: | Bio-protocol LLC | ||||||||||||
ISSN: | 2331-8325 | ||||||||||||
Official Date: | 20 April 2017 | ||||||||||||
Dates: |
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Volume: | 7 | ||||||||||||
Number: | 8 | ||||||||||||
Article Number: | e2234 | ||||||||||||
Status: | Peer Reviewed | ||||||||||||
Publication Status: | Published | ||||||||||||
Copyright Holders: | Bio-protocol LLC | ||||||||||||
Date of first compliant deposit: | 25 April 2017 | ||||||||||||
Funder: | Heptares Therapeutics (Firm), Medical Research Council (Great Britain) (MRC), Wellcome Trust (London, England) | ||||||||||||
Grant number: | MRC U105197215 | ||||||||||||
RIOXX Funder/Project Grant: |
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Open Access Version: |
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