Group-Theoretic Methods in Protein Structure Determina-n

说明（由申请人提供）：通知：no - od -09-058， NIH宣布竞争性应用恢复法案基金的可用性摘要声明“蛋白质结构测定中的群体理论方法”本补充申请的重点是分析柔性蛋白质构象的集合统计。也就是说，虽然最初的拨款侧重于从各种实验模式中确定蛋白质结构的新算法，但本补充申请的目标是量化结构数据中表示的构象的可变性。这方面最明显的例子之一是核磁共振，通常报告的是构象的整体，而不是单个结构。以下具体目标将在这个高度集中的9个月的努力中解决，增加了最初的三个具体目标：具体目标4：柔性环路构象的集成统计的有效生成。在这个特定的目标中，来自机械臂运动学领域的基于刚体运动群上的卷积和协方差传播概念的新方法将被用于生成候选环路构象的集合统计，而无需显式采样构象空间。通过规避与传统采样技术相关的指数价格，我们期望在分析蛋白质环构象的方式上产生定性差异，而不仅仅是特定计算中的定量差异。作为一个具体的测试案例，我们将把这种方法应用于人类Ube2g2的结构确定。具体目标5：评估二级结构装配中环路的偏置效应。在这个特定的目标中，我们将研究环路长度/组成与由此产生的二级结构相互作用的特殊几何特征之间的关系。我们将首先考虑螺旋-环-螺旋基序中的螺旋-螺旋相互作用以及螺旋相互作用参数与定义环的参数之间的关系。换句话说，我们将检查几何特征，如交叉角和沿螺旋长度的螺旋相互接触的位置，这取决于回路参数。这是一个纯粹的计算机目标，它将涉及从蛋白质数据库中挖掘数据。具体目标6：描述第三系系综。我们将评估以下假设：蛋白质三级结构并不总是以单一构象存在，与假设单一构象的模型相比，“三级系综”成员的枚举将更好地匹配实验数据。我们将通过验证三级系综对核磁共振残余偶极耦合来验证这一假设。特别是，我们建议利用脑膜炎奈瑟菌血红素加氧酶（nmHO）作为我们产生三级系合体的能力的测试案例，这些三级系合体保持甚至提高了与NMR数据（如rdc）的一致性。由于最初的资助目前是免费延期的，而且几乎所有的资金都已经花光了，这次竞争性的续期将为PI/co-PI的工资、一名博士后和两名学生提供一个持续九个月的资金来源。这笔费用连同拟议的实验室用品预算构成拟议的请求。

DESCRIPTION (provided by applicant): Notice: NOT-OD-09-058, NIH Announces the Availability of Recovery Act Funds for Competitive Applications SUMMARY STATEMENT "Group Theoretic Methods in Protein Structure Determination" The emphasis of this supplemental application is to analyze the statistics of ensembles of conformations of flexible proteins. That is, whereas the original grant focuses on new algorithms for determining protein structures from a variety of experimental modalities, the goal of this supplemental application is to quantify the variability of conformations represented in structural data. One of the most obvious examples of this is in NMR, where ensembles of conformations are often reported rather than an individual structure. The following specific aims to be addressed in this highly focused nine-month effort add to the original three specific aims: Specific Aim 4: Efficient Generation of Ensemble Statistics of Flexible Loop Conformations. In this specific aim, new methods from the field of robotic manipulator kinematics based on the concept of convolution and covariance propagation on the group of rigid-body motions will be used to generate ensemble statistics of candidate loop conformations without explicitly sampling conformational space. By circumventing the exponential price associated with traditional sampling techniques, we anticipate making a qualitative difference in the way protein-loop conformations are analyzed, not just a quantitative difference in particular computations. As a concrete test case, we will apply this methodology to the structure determination of human Ube2g2. Specific Aim 5: Evaluation of the Biasing Effects of Loops in the Assembly of Secondary Structures. In this specific aim we will study the correlation between loop length/composition and the special geometric characteristics of the resulting secondary structure interactions. We will begin by considering the helix-helix interaction in the helix-loop-helix motif and the relationship between helix interaction parameters and the parameters defining the loop. In other words, we will examine geometric characteristics such as crossing angle and where along the helix lengths the helices contact each other that depends on loop parameters. This is a purely in- silico objective, which will involve mining data from the Protein Data Bank. Specific Aim 6: Characterizing the Tertiary Ensemble. We will evaluate the following hypothesis: Protein tertiary structures do not always exist as single conformations, and enumeration of members of the 'tertiary ensemble' will better match experimental data than models in which a single conformation is assumed. We will test this hypothesis by validating tertiary ensembles against NMR residual dipolar couplings. In particular, we propose to utilize Neisseria meningitidis Heme Oxygenase (nmHO) as a test case of our ability to generate tertiary ensembles which maintain, or even improve, agreement with NMR data such as RDCs. Since the original grant is currently in no-cost extension, and almost all of the funds have been spent, this competitive renewal will provide an avenue for the continued funding of PI/co-PI salaries, a postdoc, and two students for a nine-month period. This, together with the proposed budget for laboratory supplies, constitutes the proposed request. PUBLIC HEALTH RELEVANCE: The emphasis of this supplemental application is to analyze the statistics of ensembles of conformations of flexible proteins. That is, whereas the original grant focuses on new algorithms for determining protein structures from a variety of experimental modalities, the goal of this supplemental application is to quantify the variability of conformations represented in structural data.