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XB-ART-46154
PLoS One 2012 Jan 01;710:e47147. doi: 10.1371/journal.pone.0047147.
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Covalent modification of mutant rat P2X2 receptors with a thiol-reactive fluorophore allows channel activation by zinc or acidic pH without ATP.

Dellal SS , Hume RI .


Abstract
Rat P2X2 receptors open at an undetectably low rate in the absence of ATP. Furthermore, two allosteric modulators, zinc and acidic pH, cannot by themselves open these channels. We describe here the properties of a mutant receptor, K69C, before and after treatment with the thiol-reactive fluorophore Alexa Fluor 546 C(5)-maleimide (AM546). Xenopus oocytes expressing unmodified K69C were not activated under basal conditions nor by 1,000 µM ATP. AM546 treatment caused a small increase in the inward holding current which persisted on washout and control experiments demonstrated this current was due to ATP independent opening of the channels. Following AM546 treatment, zinc (100 µM) or acidic external solution (pH 6.5) elicited inward currents when applied without any exogenous ATP. In the double mutant K69C/H319K, zinc elicited much larger inward currents, while acidic pH generated outward currents. Suramin, which is an antagonist of wild type receptors, behaved as an agonist at AM546-treated K69C receptors. Several other cysteine-reactive fluorophores tested on K69C did not cause these changes. These modified receptors show promise as a tool for studying the mechanisms of P2X receptor activation.

PubMed ID: 23112811
PMC ID: PMC3480388
Article link: PLoS One
Grant support: [+]

Species referenced: Xenopus laevis
Genes referenced: dtl p2rx2 p2rx4


Article Images: [+] show captions
References [+] :
Barrera, Atomic force microscopy imaging demonstrates that P2X2 receptors are trimers but that P2X6 receptor subunits do not oligomerize. 2005, Pubmed