Binding thermodynamic characterization of human P2X1 and P2X3 purinergic receptors.
Binding thermodynamic characterization of human P2X1 and P2X3 purinergic receptors.
复制标题
人类 P2X1 和 P2X3 嘌呤能受体的结合热力学特征。
DOI：
10.1016/j.bcp.2007.10.034
复制
发表时间：
2008
影响因子：
5.8
通讯作者：
P. Borea
中科院分区：
文献类型：
--
作者：
K. Varani;A. Surprenant;F. Vincenzi;A. Tosi;S. Gessi;S. Merighi;P. Borea
本研究旨在对在HEK 293细胞中表达的人P2X1和P2X3嘌呤能受体进行结合和热力学表征。测定了著名的嘌呤能激动剂和拮抗剂与P2X1和P2X3受体结合平衡的热力学参数ΔG°、ΔH°和ΔS°(标准自由能、标准自由能和标准自由能)。在4-30℃的温度范围内，使用高亲和力的嘌呤能激动剂[~3H]αβmeATP进行的饱和结合实验表明，在所研究的两种细胞系中，只有一类亲和力在纳摩尔范围内的结合位点。亲和力随温度变化，而受体密度基本上与温度无关。对于激动剂和拮抗剂，嘌呤能受体的van‘t Hoff图在4-30℃范围内呈线性关系。其热力学参数分别为−31kJol−1≤ΔH°≤−19kJol−1和17JK−1ol−1≤ΔS°≤51JK−1ol−1或−26kJol−1≤ΔH°≤36kJol−1和59≤ΔS°≤249JK−1ol−1。这些参数的结果表明，P2X1受体不受热力学区分，激动剂和拮抗剂的结合既是焓驱动的，也是熵驱动的。P2X3受体是热力学区分的，嘌呤能激动剂结合是焓和熵驱动的，而拮抗剂结合完全是熵驱动的。对这样的热力学数据的分析使我们有可能获得关于驱动嘌呤能结合相互作用的力的性质的额外信息。这些数据可能对旨在开发新的和有效的P2X1和P2X3嘌呤能配体的药物发现计划感兴趣。
The present study was designed to perform binding and thermodynamic characterization of human P2X1and P2X3purinergic receptors expressed in HEK 293 cells. The thermodynamic parameters ΔG°, ΔH° and ΔS° (standard free energy, enthalpy and entropy) of the binding equilibrium of well-known purinergic agonists and antagonists at P2X1and P2X3receptors were determined. Saturation binding experiments, performed in the temperature range 4–30°C by using the high affinity purinergic agonist [3H]αβmeATP, revealed a single class of binding sites with an affinity value in the nanomolar range in both cell lines examined. The affinity changed with the temperature whereas receptor density was essentially independent of it. van’t Hoff plots of the purinergic receptors were linear in the range 4–30°C for agonists and antagonists. The thermodynamic parameters of the P2X1or P2X3purinergic receptors were in the ranges −31kJmol−1≤ΔH°≤−19kJmol−1and 17JK−1mol−1≤ΔS°≤51JK−1mol−1or −26kJmol−1≤ΔH°≤36kJmol−1and 59≤ΔS°≤249JK−1mol−1, respectively. The results of these parameters showed that P2X1receptors are not thermodynamically discriminated and that the binding of agonists and antagonists was both enthalpy and entropy-driven. P2X3receptors were thermodynamically discriminated and purinergic agonist binding was enthalpy and entropy-driven while antagonist binding was totally entropy-driven. The analysis of such thermodynamic data makes it possible to obtain additional information on the nature of the forces driving the purinergic binding interaction. These data could be interesting in drug discovery programs aimed at development of novel and potent P2X1and P2X3purinergic ligands.