X-ray Crystallographic Fragment Screening Core

X射线晶体碎片筛选核心

• 批准号: 10363021
• 负责人: EDWARD ARNOLD
• 金额: $ 39.4万
• 依托单位: SEATTLE CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10363021
• 关键词: ray; Crystallographic; Fragment; Screening; Core

Abstract X-ray crystallographic fragment screening (XCFS) is a powerful tool for identification of new pharmacophores for probes and drug design as well as well as for biochemical and structural dynamic characterization of the protein of interest. An XCFS Core has been established to make this powerful tool accessible to multiple HIVE Projects studying HIV-1 and related proteins/complexes in the HIVE. The XCFS Core (Arnold group) has completed three XCFS campaigns against the viral targets: HIV-1 reverse transcriptase, influenza A endonuclease, and HIV-1 integrase catalytic core domain. In each case the successful screening campaign has discovered novel hits and in the cases of HIV-1 reverse transcriptase and influenza endonuclease has found novel conformations/pockets for understanding protein dynamics and biochemistry. The XCFS Core will collaborate with the other HIVE Projects and Cores to identify crystal forms that are appropriate for XCFS and optimize them as needed. Initial projects include: HIV-1 capsid (Project 3, Sarafianos), HIV-1 integrase (Projects 1 and 4, Kvaratskhelia and Engelman), and HIV-1 reverse transcriptase with nucleic acid substrate (Project 3, Arnold). Small pilot screening will verify the usefulness of the crystal form for XCFS before proceeding to large scale screening using very high-throughput synchrotron X-ray crystallography beamlines (Specific Aim 1). A specialized beamline, XChem, at the Diamond Light Source will be used to efficiently and rapidly screen 1000-2000 small molecule fragments. The XChem facility has developed several technologies for fragment soaking, crystal handling, data collection and processing to allow for very rapid XCFS campaigns. In addition to XChem, there are new high-speed beamlines at Brookhaven National Laboratories and elsewhere capable of data-collection speeds unobtainable two years ago. The new beamline technologies allow for fragments from a large library to be screened individually, whereas previously mixtures of fragments needed to be used. The ability to screen individual compounds allows for design of novel compound libraries to take advantage of this new technology (Specific Aim 2). New fragment libraries with click-reactive and SuFEx-containing moieties will be designed for XCFS (Project 6, Olson and Sharpless). Data from XCFS will be processed and refined to ensure accurate hit identification and hits will be verified through screening of derivatives and appropriate orthogonal assays. Communication and data management between the Projects and Cores will ensure clear information transfer and follow up chemistry/studies (Specific Aim 3).

抽象的 X 射线晶体碎片筛选 (XCFS) 是鉴定新药效团的强大工具 用于探针和药物设计以及生物化学和结构动态表征 感兴趣的蛋白质。 XCFS 核心的建立是为了使这个强大的工具可供多个 HIVE 访问 研究 HIV-1 和 HIVE 中相关蛋白质/复合物的项目。 XCFS Core（Arnold 组）具有 完成了针对病毒目标的三项 XCFS 活动：HIV-1 逆转录酶、甲型流感 核酸内切酶和 HIV-1 整合酶催化核心结构域。在每一个案例中，筛选活动都取得了成功 发现了新的打击，在 HIV-1 逆转录酶和流感核酸内切酶的情况下 发现了用于理解蛋白质动力学和生物化学的新构象/口袋。 XCFS 核心将与其他 HIVE 项目和核心合作，以确定晶体形式 适用于 XCFS 并根据需要对其进行优化。初始项目包括：HIV-1 衣壳（项目 3、 Sarafianos）、HIV-1 整合酶（项目 1 和 4、Kvaratskhelia 和 Engelman）和 HIV-1 逆转 带有核酸底物的转录酶（项目 3，Arnold）。小型试点筛选将验证其有用性 XCFS 的晶型，然后使用极高通量同步加速器进行大规模筛选 X 射线晶体学光束线（具体目标 1）。 Diamond Light 的专用光束线 XChem Source将用于高效、快速地筛选1000-2000个小分子片段。 XChem 设施 开发了多种碎片浸泡、晶体处理、数据收集和处理技术 允许非常快速的 XCFS 活动。除了 XChem 之外，还有新的高速光束线 布鲁克海文国家实验室和其他地方的数据收集速度在两年内是无法达到的 前。新的光束线技术允许对大型文库中的片段进行单独筛选， 而以前需要使用片段的混合物。筛选单个化合物的能力 允许设计新颖的化合物库以利用这项新技术（具体目标 2）。新的 具有点击反应性和包含 SuFEx 的片段库将为 XCFS 设计（项目 6， 奥尔森和夏普莱斯）。 来自 XCFS 的数据将被处理和细化，以确保准确的命中识别和命中验证 通过筛选衍生物和适当的正交测定。通讯和数据管理 项目和核心之间的沟通将确保清晰的信息传递和后续化学/研究（具体 目标3）。