X-ray Crystallographic Fragment Screening Core

摘要 X射线晶体学片段筛选（XCFS）是鉴定新药效团的有力工具 用于探针和药物设计以及用于生物化学和结构动态表征， 感兴趣的蛋白质一个XCFS核心已经建立，使这个强大的工具可以访问多个HIVE 研究HIV-1和相关蛋白质/复合物的项目。XCFS核心（Arnold集团） 完成了三个针对病毒目标的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。在每种情况下，成功的筛查活动 已经发现了新的命中，并在艾滋病毒-1逆转录酶和流感核酸内切酶的情况下， 发现了新的构象/口袋理解蛋白质动力学和生物化学。 XCFS核心将与其他HIVE项目和核心合作，以确定晶体形式， 并根据需要进行优化。最初的项目包括：HIV-1衣壳（项目3， Sarafianos）、HIV-1整合酶（项目1和4，Kvaratskhelia和Engelman）和HIV-1逆转录病毒（HIV-1 reverse 转录酶与核酸底物（项目3，Arnold）。小规模试点筛选将验证 在使用非常高通量同步加速器进行大规模筛选之前， X射线晶体学光束线（特定目标1）。钻石之光的专用光束线XChem Source将用于高效快速筛选1000 - 2000个小分子片段。XChem设施 已经开发了几种用于碎片浸泡、晶体处理、数据收集和处理的技术， 允许非常快速的XCFS活动。除了XChem，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无法达到两年 前新的光束线技术允许对来自大型文库的片段进行单独筛选， 而以前需要使用碎片的混合物。筛选单个化合物的能力 允许设计新型化合物库以利用这项新技术（具体目标2）。新 将为XCFS设计具有点击反应性和含SuFEx部分的片段文库（项目6， Olson和Sharpless）。 来自XCFS的数据将被处理和完善，以确保准确的命中识别和命中将被验证 通过筛选衍生物和适当的正交试验。通信和数据管理 项目和核心之间的沟通将确保明确的信息传递和后续化学/研究（具体 目标3）。

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).