Application of a Small EF Hand Affinity Tag for Expression, Purification and Biophysical Studies of G Protein-Coupled Membrane Receptors

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Title: Application of a Small EF Hand Affinity Tag for Expression, Purification and Biophysical Studies of G Protein-Coupled Membrane Receptors
Authors: Yeliseev, Alexei A.
O'Connell, David J.
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Date: 27-Feb-2019
Online since: 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.
Funding Details: Science Foundation Ireland
Type of material: Journal Article
Publisher: Russell Publishing LLC
Journal: American Pharmaceutical Review
Volume: 22
Issue: 1
Start page: 45
End page: 49
Keywords: G protein-coupled receptors (GPCR)Cell signaling pathwaysDrugs for inflammationImmunological disorders
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Language: en
Status of Item: Peer reviewed
Appears in Collections:Conway Institute Research Collection
Biomolecular and Biomedical Science Research Collection

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