使用多个重组蛋白质域进行光亲和标记位移测定
David J Fallon1,2, Alex Phillipou3, Christopher J Schofield4
1Department of Chemical Biology, GSK R&D, Gunnels Wood Road, Stevenage SG1 2NY, U.K.
The Biochemical journal
|July 4, 2023
概括
开发了一种新的光亲和标记 (PAL) 位移试验,以高效地测量化合物与odomain和外部终端域 (BET) 蛋白质的结合亲和力. 这种可访问的平台与现有方法有很强的相关性,为药物发现提供了有价值的工具.
科学领域:
- 生物化学 生物化学
- 化学生物学 化学生物学
- 药物发现 药物发现 药物发现
背景情况:
- 光亲和标记 (PAL) 是研究分子相互作用的强大技术.
- 原体和端外域 (BET) 蛋白质是关键的表观遗传调节剂和治疗点.
- 准确测量化合物结合亲和力对于药物开发至关重要.
研究的目的:
- 开发和优化一个光亲和标记 (PAL) 位移试验.
- 评估化合物与特定蛋白质结合部位的结合亲和力.
- 使用已知的BET抑制剂进行对比测试.
主要方法:
- 使用高效的PAL探头进行位移测试.
- 采用BRD4的N端和C端原蛋白作为标蛋白.
- 测试了来自ChEMBL的已知BET家族活性的264种化合物.
- 与正交时间解析的Förster共振能量转移 (TR-FRET) 数据相关的测试结果.
主要成果:
- 开发的PAL位移试验成功报告了相对结合亲缘关系.
- 用一组多样化的264种针对BET的化合物进行了对比测试.
- 获得的pIC50值与正交的TR-FRET数据有很强的相关性.
- 证明了该试验能够同时分析多个蛋白质域的能力.
结论:
- 优化的PAL位移试验是一种强大且易于使用的生化查平台.
- 这种方法为针对BET蛋白的药物发现工作提供了可靠的结合亲和数据.
- 该测试的高吞吐量潜力使其在选复合库中具有价值.
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