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Application of a Small EF Hand Affinity Tag for Expression, Purification and Biophysical Studies of G Protein-Coupled Membrane Receptors
Author(s)
Date Issued
2019-02-27
Date Available
2019-04-24T12:44:08Z
Abstract
Heptahelical G protein-coupled receptors (GPCR) comprise a large family of integral membrane proteins involved in a wide array of cell signaling pathways. For high resolution structural studies of these receptors, multi-milligram quantities of pure and structurally unperturbed proteins are required. Purification of recombinant GPCRs typically involves their solubilization into detergent micelles followed by chromatographic purification. Because of relatively low expression levels of these recombinant receptors, it is challenging to design an efficient strategy for selective and efficient purification with high yield. Here, we describe a recently introduced purification system employing a high affinity molecular switch based on fragment complementation, with a calcium dependent capture and EDTA mediated chelation elution. This technique was successfully applied to the purification of the recombinant cannabinoid receptor CB2, a promising target for the development of drugs for inflammation, immunological disorders and pain. It is feasible that similar strategies can be successfully employed for expression and purification of other membrane protein targets.
Sponsorship
Science Foundation Ireland
Other Sponsorship
Intramural Research Program of the NIAAA
Type of Material
Journal Article
Publisher
Russell Publishing LLC
Journal
American Pharmaceutical Review
Volume
22
Issue
1
Start Page
45
End Page
49
Language
English
Status of Item
Peer reviewed
ISSN
1099-8012
This item is made available under a Creative Commons License
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