Creating a Unified RAS Structural Nomenclature to Compare the Impact of Oncogenic Mutations on KRAS, NRAS, and HRAS

摘要：RAS蛋白(KRAS、NRAS和HRAS)是研究最广泛的一组突变激活蛋白。 致癌基因。然而，我们并不完全了解RAS突变对癌症的结构性影响。截至8月 2020年，蛋白质数据库中有333个实验解决的野生型(WT)和突变的RAS结构 (PDB)，包括175个HRAS、155个KRAS和3个NRAS结构。这种不断增长的结构组合提供了一种 通过统计分析发现对RAS活动的新见解的宝贵资源，从而实现量化 生物相关性的测定。近年来，核磁共振研究和分子动力学(MD)模拟 (以实验结构为模板)表明，保守的RAS开关1(SW1)和2(SW2) 区域显示修改RAS活动的动态构象行为。中的构象变化 SW1/SW2促进调控蛋白和效应蛋白在这些区域界面上的协同结合 促进RAS蛋白从活性GTP构象到非活性GDP构象的适当转换。在……里面 此外，生化研究提供了与GTP结合的RAS同源二聚体存在的证据 激活某些信号通路所必需的。虽然我们知道RAS同源二聚化发生在 GTP结合状态，我们不知道RAS同源二聚体是否需要特定的RAS构象排列 形成和相关的蛋白质-蛋白质相互作用(PPI)事件。使这个问题更加复杂的是，没有 巩固了对RAS蛋白突变如何改变SW1/SW2构象的理解 支持或不支持RAS PPI，包括RAS同源二聚化。这项提案的目标是创建一个 统一RAS结构分类以评估致癌突变对KRAS、HRAS和NRAS的影响 配体(GTP、GDP、抑制剂或无)，以及PPI/同质二聚化。在前期工作中，我创建了一个统一的 基于SW1/SW2构象跨实验结构的RAS结构命名法 KRAS、HRAS和NRAS。我还对18,841个RAS错义突变进行了泛癌症分析 100,707名患者，为KRAS、NRAS和HRAS提供最全面的现有资源 人类肿瘤中的突变模式。在目标1中，我将比较RAS的构象和PPI首选项 突变和WT形式与实验解决的结构。接下来，在目标2中，我将预测 RAS突变型和WT型的构象和PPI偏好 所产生的结构合奏。如果我们能复制一些已知突变的影响，那么我们就能 自信地预测尚未进行实验研究的新突变的后果。正在完成 在这项提议中，我将提出统一的RAS结构命名法，包括RAS确定的影响 突变，在解决新的实验结构时，数据库将不断更新。这部作品 将作为生物资源，为未来对RAS WT和突变结构和努力进行分层的研究提供信息 创造直接以RAS蛋白为靶点的药物用于癌症治疗。

Abstract: RAS proteins (KRAS, NRAS, and HRAS) are the most extensively studied set of mutationally activated oncogenes. Yet we do not completely understand the structural impact of RAS mutations in cancer. As of August 2020, there are 333 experimentally solved wild-type (WT) and mutated RAS structures in the Protein Data Bank (PDB), comprising 175 HRAS, 155 KRAS, and 3 NRAS structures. This growing structural ensemble provides a valuable resource to discover novel insights into RAS activity through statistical analyses that enable quantitative determination of biological correlates. In recent years, NMR studies and molecular dynamic (MD) simulations (using experimental structures as templates) have shown that the conserved RAS switch 1 (SW1) and 2 (SW2) regions display dynamic conformational behaviors that modify RAS activity. Conformational changes in SW1/SW2 facilitate the concerted binding of regulator and effector proteins at these regional interfaces, in turn promoting proper switching of RAS proteins from an active GTP-conformer to an inactive GDP-conformer. In addition, biochemical studies have provided evidence for the existence of a GTP-bound RAS homodimer required for activation of certain signaling pathways. While we know that RAS homodimerization occurs in a GTP-bound state, we do not know if a specific RAS conformational arrangement is required for RAS homodimer formation and associated protein-protein interaction (PPI) events. Further complicating this issue, there is no consolidated understanding of how mutations on RAS proteins may shift SW1/SW2 conformation in ways favoring or disfavoring RAS PPIs, including RAS homodimerization. The objective of this proposal is to create a unified RAS structural classification to assess the impact of oncogenic mutations on KRAS, HRAS, and NRAS, ligands (GTP, GDP, inhibitors, or none), and PPIs/homodimerization. In preliminary work, I created a unified RAS structural nomenclature based on clustering SW1/SW2 conformations across experimental structures of KRAS, HRAS, and NRAS. I have also performed a pan-cancer analysis of 18,841 RAS missense mutations from a cohort of 100,707 patients, providing the most comprehensive existing resource for KRAS, NRAS, and HRAS mutational patterns in human tumors. In Aim 1, I will compare the conformational and PPI preferences of RAS mutated and WT forms with experimentally solved structures. Following this, in Aim 2, I will predict the conformational and PPI preferences of RAS mutated and WT forms by examining energy distributions of generated structural ensembles. If we can reproduce the effects of some known mutations, then we can confidently predict the consequence for novel mutations that have not been experimentally studied. In completing this proposal, I will present the unified RAS structural nomenclature, including the determined impact of RAS mutations, in a database that will be continually updated upon solving of new experimental structures. This work will serve as a biological resource, informing future studies stratifying RAS WT and mutated structures and efforts to create drugs directly targeting RAS proteins for cancer treatment.