Structure and Function of Arp 2/3 Complex

描述(由申请人提供)：该计划项目的长期目标是了解Arp2/3复合体在内吞和细胞运动过程中控制肌动蛋白细丝组装的分子机制。Arp2/3复合体的作用机制非常复杂，需要团队合作。我们由四个地理上分散的实验室组成的小组将X射线结晶学、电子显微镜、图像处理和分子生物学结合在一起，以解决技术上具有挑战性的问题。他正在进行的合作中的结构性见解将提供关于反应机制的重要线索，也有助于设计实验来测量动力学和热力学常数，并研究活细胞中肌动蛋白的组装。项目1(Pollard)提出了一种生化和X射线结晶学实验相结合的方法，以获得有关Arp2/3复合体的构象变化和大分子相互作用的详细结构信息。项目2(LI)提出了一项关于成核促进因子与纯化的Arp2/3复合体相互作用的生物物理研究与缺乏Arp2/3复合体亚单位的小鼠细胞的显微分析相结合的研究。项目3(Hanein)建议用电子显微镜观察纯化的Arp2/3复合体、肌动蛋白细丝分支和具有和不具有功能Arp2/3复合体的小鼠细胞中的肌动蛋白网络。项目4(Volkmann)提出了从项目1和2的光谱学实验以及项目3的电子显微照片中提取最大信息量的计算方法。

DESCRIPTION (provided by applicant): The long-term goal of this program project is to understand the molecular mechanisms that allow Arp2/3 complex to control actin filament assembly during endocytosis and cellular motility. The mechanism of action of Arp2/3 complex is so complicated that a team effort is required. Our group of four geographically dispersed laboratories combines x-ray crystallography, electron microscopy, image processing and molecular biology to address technically challenging questions. Structural insights from his ongoing collaboration will provide important clues about reaction mechanisms and also help to design experiments to measure kinetic and thermodynamic constants and to study actin assembly in live cells. Project 1 (Pollard) proposes a combination of biochemical and x-ray crystallographic experiments to obtain detailed structural information regarding conformational changes and macromolecular interactions of Arp2/3 complex. Project 2 (Li) proposes a combination of biophysical studies on interaction's of nucleation promoting factors with purified Arp2/3 complex and microscopic analysis of mouse cells lacking subunits of Arp2/3 complex. Project 3 (Hanein) proposes electron microscopy of purified Arp2/3 complex, actin filament branches and of actin networks in mouse cells with and without functional Arp2/3 complex. Project 4 (Volkmann) proposes computational methods to extract the maximal amount of information from spectroscopy experiments in projects 1 and 2 and the electron micrographs from project 3.