CONFORMATION & RECOGNITION IN MICROTUBLE DYNAMICS

结构

• 批准号: 10454151
• 负责人: Luke W Rice
• 金额: $ 34.5万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10454151
• 关键词: Adopted; Affect; Affinity; Behavior; Binding; Biochemical; Biochemistry; Cells; Chromosome Segregation; Complement; Cryoelectron Microscopy; Data; Family; Genetic Materials; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; Human; Individual; Induced Mutation; Lead; Link; Measurement; Mediating; Microtubule Polymerization; Microtubules; Modeling; Molecular; Molecular Conformation; Mutation; Outcome; Paclitaxel; Pharmaceutical Preparations; Polymerase; Polymers; Property; Recombinants; Regulation; Research; Resolution; Site; Specific qualifier value; Structure; Tubulin; Tubulin Interaction; Vinca Alkaloids; Work; Yeasts; alpha Tubulin; antagonist; anti-cancer; base; beta Tubulin; design; expectation; genetic regulatory protein; insight; mutant; polymerization; reconstitution; response; segregation

Microtubules (MTs) are essential dynamic polymers required for chromosome segregation and intracellular organization, and are the direct targets of anti-cancer chemotherapeutics like taxol and the Vinca alkaloids. The dynamic properties of MTs are central to their function, and they derive from the structural and biochemical properties of individual tubulin subunits and how they interact within the MT lattice. It is increasingly appreciated that tubulin subunits adopt distinct conformations as part of the GTPase-dependent polymerization dynamics, and that regulatory proteins selectively recognize subsets of these conformations to control MT elongation, stability, and switching. The long-term goal of this research is to build a structural understanding of how allostery and the tubulin conformation cycle dictate MT dynamics, and of the mechanisms by which regulatory factors control MT dynamics. In prior project periods, we pioneered a powerful approach based on structure-inspired site-directed αβ-tubulin mutants. In the present proposal, through three specific aims, we will build on these themes to provide unique and fundamental new insights into the physical origins and regulatory mechanism of MT dynamics. We will use biochemistry, reconstitution, and modeling to define general biochemical mechanisms for XMAP215-family polymerase activity and processivity. We will reveal through structures how an `allosteric' mutation that alters MT dynamics affects tubulin conformation in human and yeast MTs, and we will provide new conformation cycle mutants to expand our understanding of allostery in MT dynamics. Finally, we will identify biochemical and structural design principles underlying how CLASP TOG interactions with tubulin suppress catastrophe and promote rescue. This work will provide new information about the conformation(s) of αβ-tubulin and how `allosteric' mutations can perturb MT dynamics and tubulin conformation. The work will also expand our understanding of how different TOG domains achieve different regulatory outcomes, with implications for the underlying mechanisms of microtubule dynamics.

微管是染色体分离和细胞内所必需的动态聚合物 组织，是抗癌化疗药物如紫杉醇和长春花碱的直接靶标。这个 MTS的动力学性质是其功能的核心，它们源于结构和生化。 单个微管蛋白亚基的性质以及它们在MT晶格中的相互作用。它越来越多地 认识到微管蛋白亚基采用不同的构象作为依赖GTPase的一部分 聚合动力学，并且调控蛋白选择性地识别这些构象的亚群以 控制MT的延伸性、稳定性和切换。这项研究的长期目标是建立一个结构性的 了解变构和微管蛋白构象周期如何决定MT动力学，以及 调控因素控制MT动态的机制。在之前的项目阶段，我们开创了一个 基于结构启发的定点定向αβ-微管蛋白突变体的有效方法。在本提案中， 通过三个具体目标，我们将以这些主题为基础，提供独特和基本的新见解 探讨MT动力学的物理起源和调控机制。我们将使用生物化学，重建， 和建模来定义XMAP215-家族聚合酶活性和 过程性。我们将通过结构揭示改变MT动力学的变构突变如何影响 微管蛋白在人和酵母MT中的构象，我们将提供新的构象周期突变体来扩展 我们对MT动力学中变构的理解。最后，我们将确定生化和结构设计 CLAP TOG如何与微管蛋白相互作用抑制灾难和促进救援的基本原理。 这项工作将提供有关αβ-微管蛋白的构象(S)和变构突变的新信息 可干扰MT动力学和微管蛋白构象。这项工作还将扩大我们对如何 不同的TOG域实现不同的监管结果，对潜在的 微管动力学机制。