ASSESSMENT OF MULTI-MICROSECOND SIMULATIONS OF INTRINSICALLY DISORDERED PROTEIN

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 这项建议的目标是将最先进的基于核磁共振的轨迹评估协议扩展到为内在无序蛋白质(IDP)计算的微秒系综。这项工作将使人们能够严格评估在最佳电流力场中编码的蛋白质动力学的时间尺度及其对其幅度的限制。该项目还将使人们能够获得高分辨率的国内流离失所者群体，目前难以在单一实验室群组上用商业软件进行计算。国内流离失所者已经成为细胞系统的关键组成部分，对细胞信号和人类疾病做出了贡献。更好地了解它们的物理化学性质是当务之急，因为IDPs的发现已经改变了科学家对蛋白质结构和功能之间关系的看法。虽然我们的内部计算基础设施足以对球状蛋白质进行简短的(&lt；1？S)模拟，但我们缺乏必要的本地资源来执行本质上无序的蛋白质的所有原子显式溶剂模拟。我要求50,000个CPU-小时，使用AMBER99SB力场计算大约40？S所有原子在人FCP1(残基930-961)的C-末端尾部显式溶剂轨迹的总模拟时间。FCP1被选为模型系统是因为它在转录终止后促进RNA聚合酶II循环的关键作用。由于我过去的成功，AMBER99SB将被用来用这个力场来模拟各种具有良好折叠蛋白质轨迹的核磁共振参数。我的实验室不断增加的内部收集的蛋白质FCP1及其与TFIIF重链蛋白质RAP74的复合体的实验核磁共振数据将有助于对使用已建立的协议计算的超长轨迹进行定量集成评估和验证。这个项目将立即给更广泛的国内流离失所者社区带来好处，因为FCP1代表了大量的国内流离失所者，他们在结合其他大分子后转变为更有序的构象。这使得这个项目成为一个理想的机会，为一个具有已知生物学意义的结合上折叠系统中涉及的所有状态生成原子系综，从而使我们能够前所未有地从机械上洞察结合事件。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The objective of this proposal is to extend state-of-the-art NMR based trajectory assessment protocols to microsecond ensembles calculated for intrinsically disordered proteins (IDPs). This work will enable a rigorous assessment of the timescale of protein dynamics encoded in the best current force fields and the restrictions to their amplitudes. This project will also allow access to high resolution ensembles of IDPs, which are currently intractable to calculate with commercial software on single-laboratory clusters. IDPs have emerged as critical components of cellular systems, contributing to cell signaling and human disease. A better understanding of their physico-chemical properties is imperative as the very discovery of IDPs has transformed scientists' view of the relationship between protein structure and function. While our in-house computational infrastructure is sufficient for short (< 1 ?s) simulations of globular proteins, we lack the local resources necessary to perform all atom explicit solvent simulations of intrinsically disordered proteins. I request 50,000 CPU-hours to calculate approximately 40 ?s total simulation time of all atom explicit solvent trajectories of the C-terminal tail of human FCP1 (residues 930-961) using the AMBER99SB force field. FCP1 is chosen as a model system because of its critical role in promoting RNA polymerase II recycling following the termination of transcription. AMBER99SB will be used due to my past success using this force field to model a variety of NMR parameters with trajectories of well folded proteins. My laboratory's ever growing body of in-house collected experimental NMR data for the protein FCP1 and its complex with the TFIIF heavy chain protein RAP74 will facilitate quantitative ensemble assessment and validation of the ultra-long trajectories computed using established protocols. This project will yield immediate benefit to the broader IDP community because FCP1 is representative of the large number of IDPs that transition to a more ordered conformation upon binding other macromolecules. This makes this project an ideal opportunity to generate atomistic ensembles for all states involved in a folding-upon-binding system with known biological significance, therefore allowing us unprecedented mechanistic insight into the binding event.