Halbfinger E, Major D T, Ritzmann M, Ubl J, Reiser G, Boyer J L, Harden K T, Fischer B
Department of Chemistry, Gonda-Goldschmied Center, Bar-Ilan University, Ramat-Gan 52900, Israel.
J Med Chem. 1999 Dec 30;42(26):5325-37. doi: 10.1021/jm990156d.
The remarkably high potencies of 2-thioether-adenine nucleotides regarding the activation of the P2Y(1)-receptor (P2Y(1)-R) in turkey erythrocyte membranes represent some of the largest substitution-promoted increases in potencies over that of a natural receptor ligand. This paper describes the investigation regarding the origin of the high potency of these P2Y(1)-R ligands over that of ATP. For this study, an integrated approach was employed combining the synthesis of new ATP analogues, their biochemical evaluation, and their SAR analysis involving NMR experiments and theoretical calculations. These experiments and calculations were performed to elucidate the conformation and to evaluate the electronic nature of the investigated P2Y(1)-R ligands. ATP analogues synthesized included derivatives where C2 or C8 positions were substituted with electron-donating groups such as ethers, thioethers, or amines. The compounds were tested for their potency to induce P2Y(1)-R-mediated activation of phospholipase C in turkey erythrocytes and Ca(2+) response in rat astrocytes. 8-Substituted ATP and AMP derivatives had little or no effect on phospholipase C or on calcium levels, whereas the corresponding 2-substituted ATP analogues potently increased the levels of inositol phosphates and ¿Ca(2+)(i). AMP analogues were ineffective except for 2-butylthio-AMP which induced a small Ca(2+) response. P2Y(1)-R activity of these compounds was demonstrated by testing these ligands also on NG108-15 neuroblastoma x glioma hybrid cells. NMR data together with theoretical calculations imply that steric, rather than electronic, effects play a major role in ligand binding to the P2Y(1)-R. Hydrophobic interactions and H-bonds of the C2 substituent appear to be important determinants of a P2Y(1)-R ligand affinity.
2-硫醚腺嘌呤核苷酸对火鸡红细胞膜中P2Y(1)受体（P2Y(1)-R）的激活具有极高的效力，相较于天然受体配体，这代表了一些因取代作用而导致效力提升最为显著的情况。本文描述了对这些P2Y(1)-R配体相较于ATP具有高效力的起源的研究。在本研究中，采用了一种综合方法，将新的ATP类似物的合成、它们的生化评估以及涉及NMR实验和理论计算的构效关系分析相结合。进行这些实验和计算是为了阐明所研究的P2Y(1)-R配体的构象并评估其电子性质。合成的ATP类似物包括C2或C8位置被供电子基团（如醚、硫醚或胺）取代的衍生物。测试了这些化合物在火鸡红细胞中诱导P2Y(1)-R介导的磷脂酶C激活以及在大鼠星形胶质细胞中引发Ca(2+)反应的效力。8-取代的ATP和AMP衍生物对磷脂酶C或钙水平几乎没有影响，而相应的2-取代的ATP类似物则显著提高了肌醇磷酸和Ca(2+)的水平。除了2-丁硫基-AMP能诱导小的Ca(2+)反应外，AMP类似物均无效。通过在NG108-15神经母细胞瘤x胶质瘤杂交细胞上测试这些配体，证实了这些化合物的P2Y(1)-R活性。NMR数据与理论计算表明，空间效应而非电子效应在配体与P2Y(1)-R的结合中起主要作用。C2取代基的疏水相互作用和氢键似乎是P2Y(1)-R配体亲和力的重要决定因素。
