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Chemically engineering ligand selectivity at the free fatty acid receptor 2 based on pharmacological variation between species orthologs

Hudson, B., Christiansen, E., Tikhonova, I.G., Grundmann, M., Kostenis, E., Adams, D.R., Ulven, T., and Milligan, G. (2012) Chemically engineering ligand selectivity at the free fatty acid receptor 2 based on pharmacological variation between species orthologs. FASEB Journal, 26 (12). pp. 4951-4965. ISSN 0892-6638 (doi:10.1096/fj.12-213314 )

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Publisher's URL: http://dx.doi.org/10.1096/fj.12-213314

Abstract

When it is difficult to develop selective ligands within a family of related G-protein-coupled receptors (GPCRs), chemically engineered receptors activated solely by synthetic ligands (RASSLs) are useful alternatives for probing receptor function. In the present work, we explored whether a RASSL of the free fatty acid receptor 2 (FFA2) could be developed on the basis of pharmacological variation between species orthologs. For this, bovine FFA2 was characterized, revealing distinct ligand selectivity compared with human FFA2. Homology modeling and mutational analysis demonstrated a single mutation in human FFA2 of C4.57G resulted in a human FFA2 receptor with ligand selectivity similar to the bovine receptor. This was exploited to generate human FFA2-RASSL by the addition of a second mutation at a known orthosteric ligand interaction site, H6.55Q. The resulting FFA2-RASSL displayed a >100-fold loss of activity to endogenous ligands, while responding to the distinct ligand sorbic acid with pEC(50) values for inhibition of cAMP, 5.83 ± 0.11; Ca(2+) mobilization, 4.63 ± 0.05; ERK phosphorylation, 5.61 ± 0.06; and dynamic mass redistribution, 5.35 ± 0.06. This FFA2-RASSL will be useful in future studies on this receptor and demonstrates that exploitation of pharmacological variation between species orthologs is a powerful method to generate novel chemically engineered GPCRs.

Item Type:Article
Status:Published
Refereed:Yes
Glasgow Author(s):Milligan, Prof Graeme and Hudson, Dr Brian
Authors: Hudson, B., Christiansen, E., Tikhonova, I.G., Grundmann, M., Kostenis, E., Adams, D.R., Ulven, T., and Milligan, G.
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:FASEB Journal
Journal Abbr.:FASEB J
ISSN:0892-6638
Published Online:23 August 2012
Copyright Holders:Copyright © 2012 The Authors
First Published:First published in FASEB Journal 26(12):4951-4965
Publisher Policy:Reproduced under a Creative Commons License
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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
589821Identification of novel FFA1 and GPR120 ligands and their use in elucidating the pharmacology, function, and clinical utility of these receptors in the treatment of obesity and diabetesBrian HudsonCanadian Institutes of Health ResearchCIHRRI MOLECULAR CELL & SYSTEMS BIOLOGY
519861Developing selective ligands for Free Fatty Acid Receptor 2 with novel pharmacological propertiesGraeme MilliganWellcome Trust (WELLCOME)089600/Z/09/ZRI NEUROSCIENCE & PSYCHOLOGY

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