RUI: Characterization and modulation of SH3 domain binding pathway biophysics

这项研究的目的是了解如何以小的方式调整常见的蛋白质结合相互作用，以在不同的细胞环境中执行专门的功能。允许细胞过程发生的细胞内通信由蛋白质之间的相互作用介导。了解这些相互作用的细节，包括它们的强度和特异性，将使研究人员能够预测和修改许多类型的细胞行为。结果将提供更深入的了解蛋白质结合相互作用如何在不同的细胞环境中发挥作用，并有助于解释一个共同的相互作用如何专门执行许多不同的细胞功能。从事该项目的本科生将有机会学习计算和实验生物物理学技能，包括计算和实验数据如何有助于项目形成更完整的蛋白质相互作用模型。学生还将与德克萨斯理工大学（Mike莱瑟姆）和利物浦大学（Elliott Stollar）的科学家密切合作，并亲身体验合作对现代科学过程的重要性。为了让更多的学生有本科研究的经验，以研究为基础的实验室课程将让学生接触到计算生物物理学和分子动力学模拟技术。在本课程中，学生开发和执行一个研究项目，为更大的项目目标做出贡献。该项目还包括为理科专业学生开设一门关于妇女和科学领域代表性不足群体的单学分课程。本课程将面向所有自然科学专业，并将鼓励学生浏览一个主要的地方，妇女传统上代表性不足，考虑和应对有关科学身份的想法。主题将包括妇女和少数民族可能面临的挑战，关于确定为科学家，刻板印象的威胁和冒名顶替者综合症。本课程将培养学生成为科学界代表性不足群体的领导者，并包括一个服务学习项目。细胞信号相互作用通常涉及内在无序的蛋白质区域与小结构域的结合，但这些相互作用的结合途径通常不太清楚。要充分理解这些无序折叠和结合相互作用如何促进功能，就需要知道如何通过调整相互作用的无序序列、结构域序列或溶剂环境来在不同背景下调整结合途径。该项目将使用SH 3结构域-肽相互作用（Abp 1 p SH 3结构域与ArkA肽结合）作为模型系统，并采用分子动力学模拟和NMR光谱相结合的方法来解决这个问题。SH 3结构域在真核生物中普遍存在，但SH 3结构域家族包含许多影响功能的变异。这些变化可以影响结合途径，这反过来又会影响生物物理特性，如结合亲和力和结合速率。这项研究将探讨SH 3-肽相互作用的不同方面如何有助于结合途径和结合的生物物理特性，最终定义一组“规则”，帮助预测这种相互作用可以通过不同的功能后果进行调节的方式。假设结合开始于ArkA和SH 3结构域之间的初始相遇，随后是ArkA片段1的快速初始结合，然后是片段2的较慢、更特异性的结合。该研究的目的1将确认两个ArkA片段在结合途径中的作用，并确定每个片段如何影响亲和力，特异性和动力学。目的2将集中在静电在结合中的作用，以及它如何影响初始转向结合位点以及其特异性结合正确的肽序列的重要性。目的3将集中于脯氨酸残基在肽序列中的重要性。脯氨酸可以通过在顺式和反式构象状态之间切换并采用刚性聚脯氨酸II螺旋结构来独特地影响构象。该项目将有助于推进我们对无序蛋白区域如何结合SH 3结构域以及不同SH 3结构域和合作伙伴之间的多样性如何有助于其功能分化的理解。了解这些生物物理性质是如何调整的，不仅会影响SH 3信号传导领域，还会影响更广泛的无序肽结合领域，因为其他识别域可能存在类似的机制。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估来支持。

The goal of this research is to understand how common protein binding interactions can be tuned in small ways to perform specialized functions in different cellular contexts. The communication within cells that allows cellular processes to occur is mediated by interactions between proteins. Understanding the details of these interactions, including their strength and specificity, will allow researchers to predict and modify the many types of cell behavior. Results will provide deeper insights into how protein binding interactions function in different cellular contexts and help explain how a common interaction can specialize to perform many different cellular functions. Undergraduate students working on this project will have the opportunity to learn both computational and experimental biophysics skills, including how both computational and experimental data can contribute to a project to form a more complete model of protein interactions. Students will also work closely with scientists at Texas Tech University (Mike Latham) and the University of Liverpool (Elliott Stollar) and experience first-hand the importance of collaboration to the modern scientific process. To allow a larger number of students to have an experience with undergraduate research, a research-based lab course will expose students to techniques in computational biophysics and molecular dynamics simulations. In this course, students develop and carryout a research project contributing to the larger project goals. This project also includes the development of a one-credit course for science majors on women and underrepresented groups in science. This course will be geared toward all natural science majors, and will encourage students navigating a major where women are traditionally underrepresented to consider and grapple with ideas about identity in science. Topics will include the challenges that women and minorities may face regarding identifying as scientists, stereotype threat, and impostor syndrome. The course will prepare students to be leaders on the topic of underrepresented groups in science and include a service-learning project.Cellular signaling interactions often involve binding of intrinsically disordered protein regions to small domains, but the binding pathways for these interactions are often not well understood. Full understanding of how these disordered folding and binding interactions contribute to function requires knowing how the binding pathway can be tuned in different contexts by adjusting the disordered sequence, domain sequence, or solvent environment of the interaction. This project will address this question using an SH3 domain-peptide interaction (Abp1p SH3 domain binding to the ArkA peptide) as a model system and employing a combination of molecular dynamics simulations and NMR spectroscopy. SH3 domains are ubiquitous across eukaryotes, but the SH3 domain family contains many variations that impact function. These variations can influence the binding pathway, which in turn will affect biophysical properties such as binding affinity and association rates. This research will investigate how different aspects of the SH3-peptide interaction contribute to the binding pathway and biophysical properties of binding, ultimately defining a set of "rules" that help predict the ways that this interaction can be modulated with different functional consequences. Binding is hypothesized to begin with an initial encounter between ArkA and the SH3 domain, followed by fast initial binding of ArkA segment 1, and then slower, more specific binding of segment 2. Aim 1 of the research will confirm the role of the two ArkA segments in the binding pathway and determine how each affects affinity, specificity, and kinetics. Aim 2 will focus on the role of electrostatics in binding, and how it affects initial steering to the binding site as well its importance for specifically binding the correct peptide sequence. Aim 3 will focus on the importance of proline residues in the peptide sequence. Proline can uniquely affect conformation by switching between cis and trans conformational states and adopting a rigid poly-proline II helix structure. The project will help to advance our understanding of how disordered protein regions bind SH3 domains and how diversity between different SH3 domains and partners contributes to their functional differentiation. Understanding how these biophysical properties are tuned will not only impact the field of SH3 signaling, but also the wider field of disordered peptide binding, as similar mechanisms may exist for other recognition domains.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.