Division of Molecular and Cellular Biosciences: Investigator-initiated research projects (MCB)

在过去的十年中，人们已经清楚地认识到，动态紊乱在蛋白质组中有着突出的表现。特别是，内在无序蛋白（IDP）广泛参与信号转导。在这个角色中，它们与它们的（结构化的）目标结合;在这样做的过程中，它们自己获得了结构秩序的度量。因此，约束过程的细节具有根本和实际的重要性。结合机制的显著特征是它通常依赖于静电相互作用。最初，IDP被长距离静电力拉向目标，形成所谓的静电遭遇复合体。从这一点开始，它迅速找到正确的构象并与目标紧密结合。为了深入了解静电相遇复合物的结构/动力学，PI将研究富含脯氨酸的肽（作为IDP的最小模型）与衔接蛋白（c-Crk N-SH 3结构域）的结合。通过将一个或两个点突变引入衔接蛋白SH 3结构域的疏水沟来改变原始系统。这就消除了紧密的束缚，并将平衡向中间状态转移。由此产生的人口增加的遭遇复杂的，使其适合于核磁共振（NMR）研究使用化学位移，顺磁约束，和弛豫数据。该实验数据集用于构建系统的计算机模型。为了实现这一目标，产生了许多~1微秒的复合物的分子动力学（MD）轨迹。这些MD数据被用作源池以形成“MD轨迹的集合”，其被定制以再现实验测量的NMR参数。值得注意的是，该模型可以严格预测固有的动态数据。这种实验校准的MD合奏有望提供一些新的见解无序蛋白质系统的结构和动力学。虽然IDP构成了一个广泛的和根本重要的一类蛋白质，他们很少或根本没有在大学教科书的覆盖面。从这个意义上说，在研究实验室和课堂教学中出现的概念之间存在着明显的差距，而且差距越来越大。PI将有助于弥合这一差距，通过（一）在几个本科院校，即布拉德利，卡尔顿，奥伯林，巴特勒和朱尼亚塔学院，所有这些都是历史上连接到普渡大学的演讲;（二）开发可供其他教师使用的视觉材料;本项目的主要研究结果将以基于MD的电影的形式呈现，由PI实验室招募的本科生设计和呈现;（iii）在《科学美国人》和/或《化学教育杂志》上发表文章，向广大受众介绍国内流离失所者。当与学生讨论IDP时，PI将强调导致蛋白质科学中出现这一新概念的“跳出框框思考”的因素。认识到积极的研究不断挑战教科书的教条，应该为感兴趣的学生提供灵感来源，并帮助引导他们中的一些人走向科学事业。特别是，PI将利用这个特殊的机会，使学生从代表性不足的群体。在这项工作中，他将依靠NOBCChE（全国黑人化学家和化学工程师专业促进组织）当地分会的持续支持。

In the last decade it has become clear that dynamic disorder is prominently represented in the proteome. In particular, intrinsically disordered proteins (IDPs) are widely involved in signal transduction. In this role, they bind to their (structured) targets; in doing so they themselves acquire a measure of structural order. The details of the binding process are, therefore, of both fundamental and practical importance. The salient feature of the binding mechanism is that it often relies on electrostatic interactions. Initially, the IDP is pulled toward its target by long-range electrostatic forces, forming what is termed an electrostatic encounter complex. Starting from this point, it quickly finds the correct conformation and binds tightly to the target. To obtain insight into the structure/dynamics of the electrostatic encounter complex, the PI will study the binding of a proline-rich peptide (which serves as a minimal model for an IDP) to an adapter protein (the c-Crk N-SH3 domain). The original system is altered by introducing one or two point mutations into hydrophobic grooves of the adapter protein SH3 domain. This abolishes tight binding and shifts the equilibrium toward the intermediate state. The resulting increase in the population of the encounter complex makes it amenable to a nuclear magnetic resonance (NMR) study using chemical shifts, paramagnetic restraints, and relaxation data. This experimental dataset is used to construct an in silico model of the system. Toward this goal, a number of ~1 micro sec molecular dynamics (MD) trajectories of the complex are generated. These MD data are used as a source pool to form an "ensemble of MD trajectories", which is tailored to reproduce the experimentally measured NMR parameters. Of note, this model can rigorously predict the pieces of data that are inherently dynamic. Such experimentally calibrated MD ensembles are expected to provide a number of new insights into the structure and dynamics of disordered protein systems. Although IDPs constitute a broad and fundamentally important class of proteins, they receive little or no coverage in college textbooks. In this sense there is a perceptible and growing gap between the concepts emerging in the research laboratories and the classroom teaching. The PI will contribute to bridging this gap by (i) making presentations at several undergraduate institutions, namely Bradley, Carleton, Oberlin, Butler, and Juniata College, all of which are historically connected to Purdue; (ii) developing visual materials that can be used by other teachers; the main findings of this project will be presented in a form of MD-based movies, designed and rendered by undergraduate students recruited into the PI's laboratory; (iii) publishing article(s) in Scientific American and/or the Journal of Chemical Education to provide a broad audience with an accessible introduction to IDPs. When discussing IDPs with students, the PI will stress the element of "thinking outside the box" that led to the emergence of this new concept in protein science. The realization that active research constantly challenges the textbook dogmas should provide a source of inspiration for interested students and help to steer some of them toward a career in science. In particular, the PI will use this special opportunity to engage the students from underrepresented groups. In this effort he will rely on the continuing support from the local chapter of NOBCChE (National Organization for the Professional Advancement of Black Chemists and Chemical Engineers).