Johnson Eric C, Feher Victoria A, Peng Jeffrey W, Moore Jonathan M, Williamson James R
Departments of Chemistry and Molecular Biology and the Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
J Am Chem Soc. 2003 Dec 24;125(51):15724-5. doi: 10.1021/ja037499s.
Several NMR screening techniques have been developed in recent years to aid in the identification of lead drug compounds. These NMR methods have traditionally been used for protein targets, and here we examine their applicability for an RNA target. We used the SHAPES compound library to test three different NMR screening methodologies: the saturation transfer difference (STD), the 2D trNOESY, and the WaterLOGSY experiments. We found that the WaterLOGSY experiment was the most sensitive method for our RNA target, the P4P6 domain of the Tetrahymena thermophila Group I intron. Using the WaterLOGSY experiment, we found that 23 of the 112 SHAPES compounds interact with P4P6. To identify which of these 23 hits bind through nonspecific interactions, we counterscreened with a linear duplex RNA control and identified one of the SHAPES compounds as interacting with P4P6 specifically. We thus demonstrated that the WaterLOGSY experiment in combination with the SHAPES compound library can be used to efficiently find RNA binding lead compounds.
近年来，已开发出几种核磁共振筛选技术，以帮助鉴定先导药物化合物。这些核磁共振方法传统上用于蛋白质靶点，在此我们研究它们对RNA靶点的适用性。我们使用SHAPES化合物库测试了三种不同的核磁共振筛选方法：饱和转移差异（STD）、二维trNOESY和WaterLOGSY实验。我们发现，对于我们的RNA靶点——嗜热四膜虫I组内含子的P4P6结构域，WaterLOGSY实验是最灵敏的方法。使用WaterLOGSY实验，我们发现112种SHAPES化合物中有23种与P4P6相互作用。为了确定这23种命中化合物中哪些是通过非特异性相互作用结合的，我们用线性双链RNA对照进行了反筛选，并确定其中一种SHAPES化合物与P4P6特异性相互作用。因此，我们证明了WaterLOGSY实验与SHAPES化合物库相结合可用于高效地找到RNA结合先导化合物。
