title: Contribution of extracellular negatively charged residues to ATP action and zinc modulation of rat P2X2 receptors.
abstract: Two histidines are known to be essential for zinc potentiation of rat P2X2 receptors, but the chemistry of zinc coordination would suggest that other residues also participate in this zinc-binding site. There is also a second lower affinity zinc-binding site in P2X2 receptors whose constituents are unknown. To assess whether the extracellular acidic residues of the P2X2 receptor contribute to zinc potentiation or inhibition, site-directed mutagenesis was used to produce alanine substitutions at each extracellular glutamate or aspartate. Two electrode voltage clamp recordings from Xenopus oocytes indicated that 7 of the 34 mutants (D82A, E85A, E91A, E115A, D136A, D209A, and D281A) were deficient in zinc potentiation and one mutant (E84A) was deficient in zinc inhibition. Additional tests on cysteine mutants at these eight positions indicated that D136 is the only residue that is a strong candidate to be at the potentiating zinc-binding site, and that E84 is unlikely to be at the inhibitory zinc-binding site.
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48105-1048, USA.
authors.forename: Sean C
authors.initials: SC
authors.lastname: Friday
authors.forename: Richard I
authors.initials: RI
authors.lastname: Hume
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology* / Q000502:physiology; D000818:Animals; D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D005110:Extracellular Space / Q000187:drug effects / Q000502:physiology*; D005260:Female; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D058476:Receptors, Purinergic P2X2; D014982:Xenopus laevis; D015032:Zinc / Q000494:pharmacology* / Q000502:physiology
chemical_list: C546403:P2rx2 protein, rat; D018048:Receptors, Purinergic P2; D058476:Receptors, Purinergic P2X2; D000255:Adenosine Triphosphate; D015032:Zinc