β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.

Ion Channel-Coupled Receptors (ICCRs) are artificial proteins comprised of a G protein-coupled receptor and a fused ion channel, engineered to couple channel gating to ligand binding. These novel biological objects have potential use in drug screening and functional characterization, in addition to...

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Autores principales:	Lydia N Caro, Christophe J Moreau, Jean Revilloud, Michel Vivaudou
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2011
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/29bc49e328c94fd691ad1071f53c93b9
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spelling	oai:doaj.org-article:29bc49e328c94fd691ad1071f53c93b92021-11-18T06:56:47Zβ2-Adrenergic ion-channel coupled receptors as conformational motion detectors.1932-620310.1371/journal.pone.0018226https://doaj.org/article/29bc49e328c94fd691ad1071f53c93b92011-03-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/21464970/?tool=EBIhttps://doaj.org/toc/1932-6203Ion Channel-Coupled Receptors (ICCRs) are artificial proteins comprised of a G protein-coupled receptor and a fused ion channel, engineered to couple channel gating to ligand binding. These novel biological objects have potential use in drug screening and functional characterization, in addition to providing new tools in the synthetic biology repertoire as synthetic K(+)-selective ligand-gated channels. The ICCR concept was previously validated with fusion proteins between the K(+) channel Kir6.2 and muscarinic M(2) or dopaminergic D(2) receptors. Here, we extend the concept to the distinct, longer β(2)-adrenergic receptor which, unlike M(2) and D(2) receptors, displayed barely detectable surface expression in our Xenopus oocyte expression system and did not couple to Kir6.2 when unmodified. Here, we show that a Kir6.2-binding protein, the N-terminal transmembrane domain of the sulfonylurea receptor, can greatly increase plasma membrane expression of β(2) constructs. We then demonstrate how engineering of both receptor and channel can produce β(2)-Kir6.2 ICCRs. Specifically, removal of 62-72 residues from the cytoplasmic C-terminus of the receptor was required to enable coupling, suggesting that ligand-dependent conformational changes do not efficiently propagate to the distal C-terminus. Characterization of the β(2) ICCRs demonstrated that full and partial agonists had the same coupling efficacy, that an inverse agonist had no effect and that the stabilizing mutation E122 W reduced agonist-induced coupling efficacy without affecting affinity. Because the ICCRs are expected to report motions of the receptor C-terminus, these results provide novel insights into the conformational dynamics of the β(2) receptor.Lydia N CaroChristophe J MoreauJean RevilloudMichel VivaudouPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 6, Iss 3, p e18226 (2011)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Lydia N Caro Christophe J Moreau Jean Revilloud Michel Vivaudou β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.
description	Ion Channel-Coupled Receptors (ICCRs) are artificial proteins comprised of a G protein-coupled receptor and a fused ion channel, engineered to couple channel gating to ligand binding. These novel biological objects have potential use in drug screening and functional characterization, in addition to providing new tools in the synthetic biology repertoire as synthetic K(+)-selective ligand-gated channels. The ICCR concept was previously validated with fusion proteins between the K(+) channel Kir6.2 and muscarinic M(2) or dopaminergic D(2) receptors. Here, we extend the concept to the distinct, longer β(2)-adrenergic receptor which, unlike M(2) and D(2) receptors, displayed barely detectable surface expression in our Xenopus oocyte expression system and did not couple to Kir6.2 when unmodified. Here, we show that a Kir6.2-binding protein, the N-terminal transmembrane domain of the sulfonylurea receptor, can greatly increase plasma membrane expression of β(2) constructs. We then demonstrate how engineering of both receptor and channel can produce β(2)-Kir6.2 ICCRs. Specifically, removal of 62-72 residues from the cytoplasmic C-terminus of the receptor was required to enable coupling, suggesting that ligand-dependent conformational changes do not efficiently propagate to the distal C-terminus. Characterization of the β(2) ICCRs demonstrated that full and partial agonists had the same coupling efficacy, that an inverse agonist had no effect and that the stabilizing mutation E122 W reduced agonist-induced coupling efficacy without affecting affinity. Because the ICCRs are expected to report motions of the receptor C-terminus, these results provide novel insights into the conformational dynamics of the β(2) receptor.
format	article
author	Lydia N Caro Christophe J Moreau Jean Revilloud Michel Vivaudou
author_facet	Lydia N Caro Christophe J Moreau Jean Revilloud Michel Vivaudou
author_sort	Lydia N Caro
title	β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.
title_short	β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.
title_full	β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.
title_fullStr	β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.
title_full_unstemmed	β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.
title_sort	β2-adrenergic ion-channel coupled receptors as conformational motion detectors.
publisher	Public Library of Science (PLoS)
publishDate	2011
url	https://doaj.org/article/29bc49e328c94fd691ad1071f53c93b9
work_keys_str_mv	AT lydiancaro b2adrenergicionchannelcoupledreceptorsasconformationalmotiondetectors AT christophejmoreau b2adrenergicionchannelcoupledreceptorsasconformationalmotiondetectors AT jeanrevilloud b2adrenergicionchannelcoupledreceptorsasconformationalmotiondetectors AT michelvivaudou b2adrenergicionchannelcoupledreceptorsasconformationalmotiondetectors
_version_	1718424156923494400

β2-Adrenergic ion-channel coupled receptors as conformational motion detectors.

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