Structure and Function of Arp 2/3 Complex

描述（由申请人提供）：本项目的长期目标是了解允许Arp2/3复合物在内吞作用和细胞运动过程中控制肌动蛋白丝组装的分子机制。Arp2/3复合物的作用机制非常复杂，需要团队的共同努力。我们的四个地理上分散的实验室组合了x射线晶体学，电子显微镜，图像处理和分子生物学来解决技术上具有挑战性的问题。从他正在进行的合作中获得的结构见解将为反应机制提供重要线索，也有助于设计实验来测量动力学和热力学常数，并研究活细胞中的肌动蛋白组装。Project 1 （Pollard）提出将生化和x射线晶体学实验相结合，获得Arp2/3复合物构象变化和大分子相互作用的详细结构信息。项目2 （Li）提出将成核促进因子与纯化Arp2/3复合物相互作用的生物物理研究与缺乏Arp2/3复合物亚基的小鼠细胞的显微分析相结合。项目3 （Hanein）提出了具有和不具有功能性Arp2/3复合物的小鼠细胞中纯化的Arp2/3复合物、肌动蛋白丝分支和肌动蛋白网络的电镜图。项目4 （Volkmann）提出了从项目1和项目2的光谱实验和项目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.