Distance-restrained docking of rifampicin and rifamycin SV to RNA polymerase using systematic FRET measurements: Developing benchmarks of model quality and reliability
Distance-restrained docking of rifampicin and rifamycin SV to RNA polymerase using systematic FRET measurements: Developing benchmarks of model quality and reliability
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DOI：
10.1529/biophysj.104.050187
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发表时间：
2005-02-01
影响因子：
3.4
通讯作者：
Levy, RM
中科院分区：
文献类型：
--
作者：
Knight, JL;Mekler, V;Levy, RM
我们正在开发用于模拟大分子复合物的距离限制对接策略，该策略结合了组件的可用高分辨率结构和源自系统荧光共振能量转移（FRET）测量的组件间距离限制。在本文中，我们考虑将小分子配体对接在大分子复合物中的问题。使用模拟的 FRET 数据，我们生成了一系列基准，允许根据 FRET 导出的距离约束的数量和质量估计模型精度，包括 FRET 导出的距离约束的数量、随机误差、系统误差、距离分布和径向分布。我们发现，对于基于以下条件的模型，预期模型精度为 10 埃或更好：i)、大于或等于 20 个约束，随机误差高达 15%，无系统误差，或 ii)、大于或等于 20 个约束，随机误差高达 15%、系统误差高达 10%，约束的对称径向分布。通过将约束数量增加到大于或等于 40 和/或通过优化约束的距离分布，可以将模型精度提高到 5 埃或更好。利用实验 FRET 数据，我们确定了小分子抑制剂利福平 (Rif) 和利福霉素 SV (Rif SV) 细菌 RNA 聚合酶内结合位点的位置。相对于晶体学定义的 Rif 结合位点，推断的 Rif 和 Rif SV 结合位点的定位精度分别为 7 埃和 10 埃。这些精度与基准模拟的预期一致，足以表明 Rif 和 Rif SV 的结合位点位于 RNA 聚合酶活性中心裂口内，与 RNA-DNA 杂交体的结合位点重叠。
We are developing distance-restrained docking strategies for modeling macromolecular complexes that combine available high-resolution structures of the components and intercomponent distance restraints derived from systematic fluorescence resonance energy transfer (FRET) measurements. In this article, we consider the problem of docking small-molecule ligands within macromolecular complexes. Using simulated FRET data, we have generated a series of benchmarks that permit estimation of model accuracy based on the quantity and quality of FRET-derived distance restraints, including the number, random error, systematic error, distance distribution, and radial distribution of FRET-derived distance restraints. We find that expected model accuracy is 10 Angstrom or better for models based on: i), greater than or equal to20 restraints with up to 15% random error and no systematic error, or ii), greater than or equal to20 restraints with up to 15% random error, up to 10% systematic error, and a symmetric radial distribution of restraints. Model accuracies can be improved to 5 Angstrom or better by increasing the number of restraints to greater than or equal to40 and/or by optimizing the distance distribution of restraints. Using experimental FRET data, we have defined the positions of the binding sites within bacterial RNA polymerase of the small-molecule inhibitors rifampicin (Rif) and rifamycin SV (Rif SV). The inferred binding sites for Rif and Rif SV were located with accuracies of, respectively, 7 and 10 Angstrom relative to the crystallographically defined binding site for Rif. These accuracies agree with expectations from the benchmark simulations and suffice to indicate that the binding sites for Rif and Rif SV are located within the RNA polymerase active-center cleft, overlapping the binding site for the RNA-DNA hybrid.