Studies on Microtubule Binding Proteins

描述(申请人提供)：肌动蛋白沿微管移动或从末端解聚。在过去的12年里，我们使用冷冻电子显微镜和图像分析来研究这些运动和解聚活性。基于我们的发现，以及许多其他实验室的工作，我们概述了统一我们对动蛋白和肌球蛋白的想法的设计原则，并为从机制上理解这两个分子马达超家族提供了一个总体框架。我们现在已经对核苷酸介导的构象变化有了很好的总体理解，包括传统的动蛋白正端定向运动，14s动蛋白的非进行性和负端定向运动，以及动蛋白13介导的解聚的许多方面。对微管相关蛋白的更多研究为这些蛋白选择性稳定微管的机制提供了洞察力。我们过去的结果为当前的提议奠定了基础。这里描述的实验分为两个大的主题领域。首先，我们描述了一些实验，这些实验将回答我们认为剩余的一些关于动蛋白介导的运动性和解聚的悬而未决的一般性问题。其次，我们在MAP研究的基础上，扩展了我们研究计划的这一方面，并纳入了对微管+末端跟踪蛋白和动粒复合体的研究，以探讨微管动力学和染色体-微管连接的调控的结构基础。在我们所有的工作中，我们寻求回答基本的生物学问题，我们的发现与健康和疾病状态都相关。

DESCRIPTION (provided by applicant): Kinesins move along microtubules or depolymerize them from the ends. Over the last 12 years, we have used cryo-electron microscopy and image analysis to investigate these motility and depolymerization activities. Based on our findings, as well as work from many other laboratories, we have outlined design principles that unify our thinking about kinesins and myosins and provide a general framework for a mechanistic understanding of these two superfamilies of molecular motors. We now have a good general understanding of the nucleotide-mediated conformational changes underlying processive, plus-end-directed motion by conventional kinesin, non-processive, minus-end-directed motion by kinesin 14s, and many aspects of kinesin 13-mediated depolymerization. Additional work on microtubule associated proteins has provided insights into the mechanisms of selective microtubule stabilization by these proteins. Our past results set the stage for the current proposal. The experiments described here fall into two broad subject areas. First, we describe experiments that will answer what we see as some of the remaining outstanding general questions about kinesin-mediated motility and depolymerization. Second, we build upon our investigations on MAPs, expanding this aspect of our research program and incorporating studies on microtubule plus-end-tracking proteins and kinetochore complexes to investigate the structural basis for modulation of microtubule dynamics and chromosome-microtubule attachments. In all our work, we seek to answer fundamental biological questions and our findings have relevance for both the healthy and diseased states.