RUI: Biophysical investigation of SH3 domain binding partners: How the binding motif and surrounding disordered sequence affect the finding pathway

细胞内允许细胞过程发生的通信是通过蛋白质之间的相互作用来调节的。这项研究的目标是了解这些基本的相互作用是如何在相互作用的地方以及周围地区受到控制的。了解这些相互作用的细节将使研究人员能够预测和修改细胞行为。该项目的结果将提供对蛋白质结合相互作用在不同环境中如何发挥作用的更深入的见解，包括在存在可以结合同一蛋白质伙伴的多个位点的情况下，并有助于解释常见的相互作用如何专门化来执行许多不同的细胞功能。参与这个项目的本科生将有机会学习计算和实验生物物理学技能。为了让更多的学生从事本科研究，研究人员将提供一门以研究为基础的实验室课程，让学生接触计算生物物理学和分子动力学模拟的技术。在本课程中，学生将开发并完成一个对更大的项目目标做出贡献的研究项目。研究人员还将开设一门为期两个学分的课程，内容是关于科学中的身份认同，该课程将鼓励自然科学专业的学生考虑并努力思考关于科学身份的想法，以及如何让科学更具包容性。学生将创建具体的干预措施，以改善未被充分代表的学生在科学领域的体验。本课程将培养学生成为科学中同一性和包容性这一主题的领导者。细胞信号相互作用经常涉及到内在无序的蛋白质区域与小区域的结合。这些相互作用的亲和力和特异性取决于无序区域内的结合基序，但也受到侧翼区域和周围环境的影响。此外，结合途径，而不仅仅是结合状态的结构，对于理解这些相互作用的功能适应可能是重要的。以SH3结构域结合为模型系统，PI已经用分子动力学(MD)模拟表明无序的富含Pro的多肽Arka通过多步过程与AbpSH3结构域结合。在这个项目中，PI现在将通过使用MD模拟、核磁共振光谱和ITC在日益复杂的水平上系统地研究这一结合途径，来研究这一结合途径在不同生物相关环境中的变化。第一个目标将集中在通过模拟与顺式构象中的Arka Pro结合、盐屏蔽静电相互作用以及参与结合的带电AbpSH3残基的突变来改变结合基序本身的柔性和静电相互作用的影响。在第二个目的中，将比较不同的生理相关的IDR序列，以探索基序侧翼区对结合的影响。在第三个目标中，将研究多个结合基序背景下的结合过程。该项目的完成将把无序蛋白质区域如何与SH3结构域结合的理解增加到一个新的复杂性水平，因为结合基序和周围序列如何影响结合途径，最终将揭示生物功能。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The communication within cells that allows cellular processes to occur is mediated by interactions between proteins. The goal of this research is to understand how these fundamental interactions are controlled at the local site of interaction, as well as the surrounding regions. Understanding the details of these interactions would allow researchers to predict and modify cell behavior. Results of this project will provide deeper insights into how protein binding interactions function in different contexts, including in the presence of multiple sites that can bind the same protein partner, 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. To allow a larger number of students to engage in undergraduate research, the investigator will offer a research-based lab course that will expose students to techniques in computational biophysics and molecular dynamics simulations. In this course, students will develop and carryout a research project contributing to the larger project goals. The investigator will also offer a two-credit course on Minoritized Identities in Science that will encourage natural science majors to consider and grapple with ideas about identity in science and how to make the sciences more inclusive. Students will create specific interventions to improve the experience of underrepresented students in science. The course will prepare students to be leaders on the topic of identity and inclusivity in science.Cellular signaling interactions often involve binding of intrinsically disordered protein regions to small domains. The affinity and specificity of these interactions depend on the binding motif within the disordered region, but are also affected by the flanking regions and surrounding context. Additionally, the binding pathway, rather than just the structure of the bound state, can be important for understanding the functional adaptation of these interactions. Focusing on SH3 domain binding as a model system, the PI have previously shown that the disordered proline-rich peptide ArkA binds to the AbpSH3 domain in a multi-step process using molecular dynamics (MD) simulations. In this project, the PI will now examine how this binding pathway varies in different biologically relevant contexts by systematically studying this pathway at increasing levels of complexity using MD simulations, NMR spectroscopy, and ITC. The first aim will focus on the effects of altering the flexibility and electrostatic interactions of the binding motif itself by simulating the binding with ArkA prolines in the cis conformation, with salt screening electrostatic interactions, and with mutations to charged AbpSH3 residues involved in binding. In the second aim, different physiologically relevant IDR sequences will be compared to explore the effect of the motif-flanking regions on binding. In the third aim, the binding process in the context of multiple binding motifs will be examined. The completion of this project will add a new level of complexity to the understanding of how disordered protein regions bind to SH3 domains, as how both the binding motif and the surrounding sequences influence the binding pathway, and ultimately biological function will be revealed.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.