Computational Studies of Model Systems for Protein Unfolded States

虽然蛋白质的未折叠状态以前被认为在很大程度上是随机的，但特异性的证据表明， 结构在积累。即使在低种群中，这种类型的排序也可能对蛋白质产生直接影响。 稳定性、折叠机制和速率，以及在许多细胞中涉及的错误折叠和聚集， 人类疾病在大多数情况下，弱性质的残余结构和低人口的错误折叠 在生理条件下的状态排除了使用 实验技术模拟已经开始为实验提供补充数据，因为它们 可以在大多数人无法达到的时间尺度上提供单分子分辨率的结构细节， 实验然而，由于多种原因，模拟在许多重要情况下失败。 该提案概述了模拟算法和力场的持续发展， 应用于研究展开状态，直接验证实验数据，通过 建立合作。一个强烈的重点放在构象采样;一个主要组成部分， 该项目涉及开发一种新的采样方法， 系综数据与显着降低计算成本的溶剂。 模拟将在不同复杂性的几个模型系统上进行。在每种情况下，具体 突变将探测假设参与决定结构或稳定性的相互作用 的原生褶皱。其他突变体探测我们在实验中观察到的相互作用稳定性的影响。 展开状态。这些研究将为这些重要的模型系统提供有用的见解， 对我们的力场和采样方法的性能提供宝贵和关键的反馈。 研究的下一阶段涉及每个模型系统的展开集合的表征， 产生关于任何残余结构的性质的实验可检验的假设。中的每 模型系统，最近的实验表明，残余结构可能存在于未折叠状态，我们的 模拟将为解释这些数据提供重要的模型。展开的其他研究 状态和熵在折叠中的作用将通过柔性甘氨酸和刚性甘氨酸的替换来实现。 脯氨酸与丙氨酸的非折叠态熵和折叠自由能的影响。

While unfolded states of proteins were previously thought to be largely random, evidence of specific structure is accumulating. Even at low populations this type of ordering could have direct bearing on protein stability, folding mechanism, and rate, as well as the misfolding and aggregation that are implicated in many human disorders. In most cases, the weak nature of the residual structure and low populations of misfolded states under physiological conditions preclude direct and detailed characterization of these ensembles using experimental techniques. Simulations have begun to provide data complementary to experiments, since they can provide structural detail with single molecule resolution on a time scale inaccessible to most experiments. However, simulations fail in many important cases for multiple reasons. This proposal outlines continued development of simulation algorithms and force fields to enable successful application to the study of unfolded states, with direct validation against experimental data obtained through established collaborations. A strong emphasis is placed on conformational sampling; a major component of the project involves development of a novel sampling approach that provides improved convergence of ensemble data with explicit inclusion of solvent at a significantly reduced computational cost. Simulations will be performed on several model systems of varying complexity. In each case, specific mutations will probe interactions that are hypothesized to be involved in determining the structure or stability of the native fold. Other mutants probe the effect on stability of interactions that we have observed in the unfolded state. These studies will provide useful insight into these important model systems, and will also provide valuable and critical feedback on the performance of our force fields and sampling methods. The next phase of the research involves characterization of unfolded ensembles for each model system, and generation of experimentally testable hypotheses about the nature of any residual structure. For each of the model systems, recent experiments suggest that residual structure may exist in the unfolded state, and our simulations will provide important models for interpretation of this data. Additional studies of the unfolded state and the role of entropy in folding will be performedthrough replacement of flexible glycine and rigid proline with alanine, investigating the effect on unfolded state entropy and free energy of folding.