Nakazawa K, Ohno Y, Inoue K
Division of Pharmacology, National Institute of Health Sciences, Tokyo, Japan.
Biochem Biophys Res Commun. 1998 Mar 17;244(2):599-603. doi: 10.1006/bbrc.1998.8312.
Charged or polarized amino acid residues near or within the second transmembrane (M2) segment of neuronal ATP receptor/channels (P2X2 receptors) were neutralized by site-directed mutagenesis, and the properties of the mutants were electrophysiologically characterized using Xenopus oocytes. When Asp315 was substituted with Val (D315V), the sensitivity to ATP was reduced by about 60-fold. The sensitivity to ATP was not affected by the neutralization of Lys324, which is involved in a Walker type A ATP-binding sequence, Lys366, Tyr330, or Asn333. With D315V channels, the sensitivities to other agonists (ADP, ATP gamma S, and 2-methylthio ATP) were also reduced. The sensitivities to antagonists (suramin and Cibacron Blue F3GA) were, however, not affected by this neutralization. The results suggest that Asp315, which is assumed to be present in the extracellular region near the M2 segment of P2X2 receptor/channels, serves to maintain agonist sensitivity.
通过定点诱变中和了神经元ATP受体/通道（P2X2受体）第二个跨膜（M2）片段附近或内部的带电荷或极化的氨基酸残基，并使用非洲爪蟾卵母细胞对突变体的特性进行了电生理学表征。当用缬氨酸取代天冬氨酸315（D315V）时，对ATP的敏感性降低了约60倍。对ATP的敏感性不受参与沃克A型ATP结合序列的赖氨酸324、赖氨酸366、酪氨酸330或天冬酰胺333中和的影响。对于D315V通道，对其他激动剂（ADP、ATPγS和2-甲硫基ATP）的敏感性也降低了。然而，对拮抗剂（苏拉明和汽巴蓝F3GA）的敏感性不受这种中和的影响。结果表明，假定存在于P2X2受体/通道M2片段附近细胞外区域的天冬氨酸315有助于维持激动剂敏感性。
