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In P2X receptors, agonist binding at the interface between neighbouring subunits is efficiently transduced to ion channel gating. However, the relationship between binding and gating is difficult to study because agonists continuously bind and unbind. Here, we covalently incorporated agonists in the binding pocket of P2X receptors and examined how binding site occupancy affects the ability of the channel to gate.We used a strategy for tethering agonists to their ATP-binding pocket, while simultaneously probing ion channel gating using electrophysiology. The agonist 2',3'-O-(4-benzoylbenzoyl)-ATP (BzATP), a photoaffinity analogue of ATP, enabled us to trap rat homomeric P2X2 receptor and a P2X2/1 receptor chimera in different agonist-bound states. UV light was used to control the degree of covalent occupancy of the receptors.Irradiation of the P2X2/1 receptor chimera - BzATP complex resulted in a persistent current that lasted even after extensive washout, consistent with photochemical tethering of the agonist BzATP and trapping of the receptors in an open state. Partial labelling with BzATP primed subsequent agonist binding and modulated gating efficiency for both full and partial agonists.Our photolabelling strategy provides new molecular insights into the activation mechanism of the P2X receptor. We show here that priming with full agonist molecules leads to an increase in gating efficiency after subsequent agonist binding.
Agboh,
Characterisation of ATP analogues to cross-link and label P2X receptors.
2009, Pubmed
Agboh,
Characterisation of ATP analogues to cross-link and label P2X receptors.
2009,
Pubmed Aschrafi,
Trimeric architecture of homomeric P2X2 and heteromeric P2X1+2 receptor subtypes.
2004,
Pubmed
,
Xenbase Barrera,
Atomic force microscopy imaging demonstrates that P2X2 receptors are trimers but that P2X6 receptor subunits do not oligomerize.
2005,
Pubmed Biskup,
Relating ligand binding to activation gating in CNGA2 channels.
2007,
Pubmed
,
Xenbase Bongartz,
Aminoglycoside block of P2X2 receptors heterologously expressed in Xenopus laevis oocytes.
2010,
Pubmed
,
Xenbase Brown,
Specific labeling and permanent activation of the retinal rod cGMP-activated channel by the photoaffinity analog 8-p-azidophenacylthio-cGMP.
1993,
Pubmed Ding,
Single channel properties of P2X2 purinoceptors.
1999,
Pubmed
,
Xenbase Erb,
Covalent incorporation of 3'-O-(4-benzoyl)benzoyl-ATP into a P2 purinoceptor in transformed mouse fibroblasts.
1990,
Pubmed Evans,
Orthosteric and allosteric binding sites of P2X receptors.
2009,
Pubmed Forman,
Photoactivated 3-azioctanol irreversibly desensitizes muscle nicotinic ACh receptors via interactions at alphaE262.
2007,
Pubmed
,
Xenbase Jiang,
Agonist trapped in ATP-binding sites of the P2X2 receptor.
2011,
Pubmed Kapfer,
Comparative photoaffinity labeling study between azidophenyl, difluoroazidophenyl, and tetrafluoroazidophenyl derivatives for the GABA-gated chloride channels.
1995,
Pubmed Karpen,
Covalent activation of retinal rod cGMP-gated channels reveals a functional heterogeneity in the ligand binding sites.
1996,
Pubmed
,
Xenbase Karpen,
Ion channels: does each subunit do something on its own?
2002,
Pubmed Kawate,
Crystal structure of the ATP-gated P2X(4) ion channel in the closed state.
2009,
Pubmed Khakh,
P2X receptors as cell-surface ATP sensors in health and disease.
2006,
Pubmed Kilkenny,
Animal research: reporting in vivo experiments: the ARRIVE guidelines.
2010,
Pubmed Le Novère,
The Ligand Gated Ion Channel Database.
1999,
Pubmed Li,
Gating the pore of P2X receptor channels.
2008,
Pubmed Marquez-Klaka,
Identification of an intersubunit cross-link between substituted cysteine residues located in the putative ATP binding site of the P2X1 receptor.
2007,
Pubmed
,
Xenbase McGrath,
Guidelines for reporting experiments involving animals: the ARRIVE guidelines.
2010,
Pubmed Mourot,
Agonist- and competitive antagonist-induced movement of loop 5 on the alpha subunit of the neuronal alpha4beta4 nicotinic acetylcholine receptor.
2008,
Pubmed
,
Xenbase Mourot,
Photo-induced covalent attachment of agonists as a tool to study allosteric mechanisms of nicotinic acetylcholine receptors.
2006,
Pubmed Nicke,
P2X1 and P2X3 receptors form stable trimers: a novel structural motif of ligand-gated ion channels.
1998,
Pubmed
,
Xenbase North,
Pharmacology of cloned P2X receptors.
2000,
Pubmed North,
Molecular physiology of P2X receptors.
2002,
Pubmed Rettinger,
Desensitization masks nanomolar potency of ATP for the P2X1 receptor.
2004,
Pubmed
,
Xenbase Rettinger,
Activation and desensitization of the recombinant P2X1 receptor at nanomolar ATP concentrations.
2003,
Pubmed
,
Xenbase Ruiz,
Single cyclic nucleotide-gated channels locked in different ligand-bound states.
1997,
Pubmed
,
Xenbase Valera,
A new class of ligand-gated ion channel defined by P2x receptor for extracellular ATP.
1994,
Pubmed
,
Xenbase Virginio,
Trinitrophenyl-substituted nucleotides are potent antagonists selective for P2X1, P2X3, and heteromeric P2X2/3 receptors.
1998,
Pubmed Werner,
Domains of P2X receptors involved in desensitization.
1996,
Pubmed
,
Xenbase Zhong,
Photo-activated azi-etomidate, a general anesthetic photolabel, irreversibly enhances gating and desensitization of gamma-aminobutyric acid type A receptors.
2008,
Pubmed
,
Xenbase
