RUI: D3SC: The Role of Non-Motif Selectivity Determinants in Peptide-Binding Domain Interactions

细胞接收并响应局部信号以与环境沟通，这些细胞信号传导途径中的错误可导致许多不同的人类疾病。描述这种和其他细胞途径的一个重大挑战是，信号蛋白之间的许多相互作用仅依赖于氨基酸构建块之间的少量接触，导致结合伙伴的乱交和重叠。凭借这一奖项，化学部门的生命过程化学项目资助了西华盛顿大学的Jeanine Amacher博士，研究细胞信号蛋白如何在它们之间的接触数量限制在10个或更少的氨基酸的情况下特异性地识别彼此。本项目描述了细胞信号通路中每一个参与蛋白质识别的相互作用氨基酸，并阐明了这些相互作用是如何进化的。此外，该项目还为本科生和硕士研究生提供结构生物学和蛋白质生物化学的第一手经验，为他们从事stem相关的职业做好准备。细胞中一类重要的蛋白质-蛋白质相互作用涉及对短线性基序（slms）或肽的识别。这些相互作用对细胞中的信号传导和运输成分至关重要，但通常在亲和力和短暂性方面相对较弱。例子包括激酶、磷酸酶、SH2和PDZ结构域介导的相互作用。然而，slim结合结构域仅识别几个氨基酸位置导致相互作用网络大大重叠，使得仅基于基序分析的解剖生物学上重要的相互作用几乎是不可能的。在这个项目中，PDZ结构域被用作一个模型系统来定义非基序选择性决定因素在SLiM或肽介导的相互作用中的作用。PDZ结构域通过仅基于两个残基的结合基识别靶蛋白的极端c端。同样地，两个序列可以结合数量截然不同的PDZ结构域。例如，人乳头瘤病毒E6癌蛋白（HPV16 E6）的C端序列与十几种含PDZ结构域的蛋白相互作用，而囊性纤维化跨膜传导调节因子（CFTR）的C端序列与不到五种蛋白相互作用。hpv16e6和CFTR都含有相同的基序残基。该项目的总体目标是：(i)剖析这两个目标序列在非基序或调制器残基上的混杂性，以及（ii）定义调制器残基在整个进化过程中的作用。这个提议的中心假设是，调制器的偏好决定了细胞的相互作用网络。从这个项目中产生的原则适用于其他slim绑定域族。该奖项反映了美国国家科学基金会的法定使命，并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，被认为值得支持。

Cells receive and respond to local signals to communicate with their environment, and errors in these cell signaling pathways can lead to a number of different human diseases. A significant challenge in describing this and other cellular pathways is that many interactions among signaling proteins rely on just a small number of contacts between their amino acid building blocks, leading to promiscuity and overlap in binding partners. With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Jeanine Amacher from Western Washington University to investigate how cell signaling proteins can specifically recognize one another with the number of contacts limited to 10 or less amino acids between them. This project characterizes every interacting amino acid involved in protein recognition for one set of proteins along the cell signaling pathway and elucidates how these interactions evolved. In addition, the project provides undergraduate and masters level graduate students with firsthand experience in structural biology and protein biochemistry to prepare them for STEM-related careers. An important class of protein-protein interactions in the cell involve recognition of short linear motifs (SLiMs) or peptides. These interactions are critical in signaling and trafficking components in the cell, but are often relatively weak in affinity and transient. Examples include kinase, phosphatase, SH2, and PDZ domain-mediated interactions. However, the recognition of only a couple of amino acid positions by SLiM-binding domains results in interaction networks that greatly overlap, rendering the dissection biologically-important interactions nearly impossible based on the analysis of motifs alone. In this project, the PDZ domain is used as a model system to define the role of non-motif selectivity determinants in SLiM- or peptide mediated interactions. PDZ domains recognize the extreme C-terminus of target proteins through binding motifs that are based on only two residues. Likewise, two sequences can bind dramatically different numbers of PDZ domains. For example, the C-terminal sequence of the human papillomavirus E6 oncoprotein (HPV16 E6) interacts with over a dozen PDZ domain containing proteins, while the C terminal sequence of cystic fibrosis transmembrane conductance regulator (CFTR) interacts with less than five. Both HPV16 E6 and CFTR contain identical motif residues. The overall objectives in this project are to (i) dissect the promiscuity of these two target sequences at non-motif, or modulator, residues and (ii) define the role of modulator residues throughout evolution. The central hypothesis of this proposal is that modulator preferences dictate cellular interaction networks. The principles emerging from this project are applicable to other SLiM-binding domain families.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.