Allosteric elements in the superfamily of small GTPases

小GTP酶是参与细胞功能的几乎所有方面的蛋白质，从控制细胞增殖、调节细胞死亡、运动、在细胞内运输含有分子货物的膜小泡以及在细胞质和细胞核之间运输分子。它们的活性受与其活性部位结合的分子(GTP)转化为GDP和无机磷的调节。它们属于Ras、Rho、Rab、Arf和Ran 5个家族的150多种蛋白质。这项研究的目标是确定在细胞膜上发现的具有代表性的Ras、Rho和Arf家族蛋白是如何进化出与其不同功能相关的特定结构细节的。研究人员将测试与细胞膜的相互作用如何影响这些小GTP酶的功能。这将使用基于互补的最先进的生物物理和生物化学实验的多学科方法来完成，旨在阐明这些蛋白质工作的基本分子机制。除了继续促进这个项目将在不同的实验室和部门环境中进行外，还将在大学层面上推出一个新的未来教师奖学金指导计划，与东北大学的高级计划相关。Ras超家族GTP酶(小的GTP酶)中的蛋白质参与细胞中非常不同的功能，它们通过一个严格调控的机制工作，从活跃的GTP结合状态转换到不活跃的GDP结合状态。现在有大量证据表明，与膜的相互作用是通过变构调节将膜相互作用的热点连接到活性部位的调节机制的组成部分。在这个项目中，研究人员的目标是确定这些相互作用的性质，并绘制超家族中三个具有代表性的成员：HRA、RhoA和Arf1的变构途径。研究人员假设，特定的膜元件在调节超家族大多数分支的G蛋白功能方面发挥着关键作用。他们将通过使用一系列磷脂头部基团模拟物来确定膜组件与HRAs、RhoA和Arf1的结合特异性，并通过研究策略性设计的突变体对状态构象集成和相关水解率的影响来验证这一假说。这将使用X射线结晶学、水解率测量、广角区域(WAXS)中的溶液散射实验和加速MD(AMD)模拟相结合的方法来完成，每种方法都提供了一个视角，这些方法一起将导致对调控机制的机械观点，这些机制在超级家族中迄今尚未被探索过。

Title: Allosteric elements in the superfamily of small GTPasesThe Small GTPases are proteins involved in virtually all aspects of cellular function, from the control of cell proliferation, regulation of cell death, motility, transport of membrane vesicles containing molecular cargo within the cell and transport of molecules between the cell cytoplasm and nucleus. Their activity is regulated by conversion of a molecule (GTP) bound to their active site to GDP and inorganic phosphate. They belong to a group of over 150 proteins classified into 5 families: Ras, Rho, Rab, Arf and Ran. The goal of this research project is to determine how representative proteins in the Ras, Rho and Arf families, protein families that are found on the cell membrane, have evolved specific structural details associated with their distinct functions. The investigators will test how interactions with the cell membrane affect the function of these small GTPases. This will be done using a multidisciplinary approach based on complementary state of the art biophysical and biochemical experiments designed to elucidate the fundamental molecular mechanisms through which these proteins work. In addition to continuing to promote diverse laboratory and departmental environments in which this project will be conducted, there will be a new Future Faculty Fellowship mentoring program at the university level associated with the ADVANCE program at Northeastern. Proteins in the Ras superfamily of GTPases (Small GTPases) are involved in vastly diverse functions in the cell, where they work through a tightly regulated mechanism of switching from an active GTP-bound state to an inactive GDP-bound state. There is now substantial evidence that interaction with the membrane is an integral part of the regulatory mechanism through allosteric modulation linking membrane-interacting hot spots to the active site. In this project, investigators aim to determine the nature of these interactions and to map the allosteric pathways in three representative members of the superfamily: HRas, RhoA and Arf1. The investigators hypothesize that specific membrane elements play a key role in regulating the function of G-proteins across most branches of the superfamily. They will will test this hypothesis by using a series of phospholipid head group mimics to determine the binding specificities of membrane components to HRas, RhoA and Arf1 and by studying the effects of strategically designed mutants on the conformational ensemble of states and associated hydrolysis rates. This will be done using a combination of X-ray crystallography, hydrolysis rate measurements, solution scattering experiments in the wide angle regime (WAXS) and accelerated MD (aMD) simulations, each method providing a perspective that together will lead to a mechanistic view of regulatory mechanisms that have thus far been unexplored within the superfamily.