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PLoS One. 2017 Nov 1;12(11):e0187306. doi: 10.1371/journal.pone.0187306. eCollection 2017.

Generation of G protein-coupled receptor antibodies differentially sensitive to conformational states.

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Proteimax Biotechnology LTDA, Av Corifeu de Azevedo Marques, São Paulo, SP, Brazil.
Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, United States of America.
Department of Pharmacology, Biomedical Science Institute, University of São Paulo, São Paulo, SP, Brazil.
Department of Pharmacology, Temple University School of Medicine, Philadelphia, United States of America.


The N-terminal region of G protein-coupled receptors can be efficiently targeted for the generation of receptor-selective antibodies. These antibodies are useful for the biochemical characterization of the receptors. In this study, we developed a set of criteria to select the optimal epitope and applied them to generate antibodies to the N-terminal region of 34 different G protein-coupled receptors. The antibody characterization revealed that a subset of antibodies exhibited increased recognition of the receptor following agonist treatment and this increase could be blocked by treatment with the receptor antagonist. An analysis of the epitopes showed that those antibodies that exhibit increased recognition are on average twelve residues long, have an overall net negative charge and are enriched in aspartic and glutamic acids. These antibodies are useful since they facilitate studies examining dose dependent increases in recognition of receptors in heterologous cells as well as in native tissue. Another interesting use of these antibodies is that they facilitate measuring changes in receptor recognition in brain following peripheral drug administration; for example, systemic administration of cocaine, a blocker of dopamine transporter that increases local dopamine levels at the synapse, was found to lead to increases in antibody recognition of dopamine receptors in the brain. Taken together these studies, in addition to describing novel tools to study native receptors, provide a framework for the generation of antibodies to G protein-coupled receptors that can detect ligand-induced conformational changes.

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