PROTEIN COMPOSITION OF ACTIN TAILS ASSEMBLED IN YEAST EXTRACTS

这个子项目是利用资源的许多研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 为子项目列出的总成本可能 代表子项目使用的中心基础设施的估计数量， NCRR赠款不直接向子项目或子项目工作人员提供资金。 网格蛋白介导的内吞作用是活细胞中的一个基本过程，使它们能够有效地从细胞外环境捕获大分子并将其包装到胞质溶胶中的囊泡中。许多疾病都可以追溯到导致内吞作用的分子反应的连续缺陷。在芽殖酵母酿酒酵母中，胞吞作用定位于皮层肌动蛋白斑块，在那里，肌动蛋白丝的密集网络聚合在网格蛋白包被的凹坑周围。该网络由一组复杂的蛋白质调节，这些蛋白质合作以产生膜内陷。 我们最近在我们的实验室开发了一个系统，在酵母提取物的组装肌动蛋白为基础的结构周围的聚苯乙烯微珠。我们相信，这些结构的蛋白质组成可能是类似的蛋白质组成中发现的皮质肌动蛋白补丁在体内。我们分离出了具有相关肌动蛋白结构的珠子，我们想通过质谱来确定蛋白质的组成。这些结果将使我们能够进一步分析不同成分在胞吞作用中的作用。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Clathrin-mediated endocytosis is a fundamental process in living cells, allowing them to capture macromolecules efficiently from the extracellular environment and package them into vesicles in the cytosol. Numerous diseases can be traced to a defect in the succession of molecular reactions leading to endocytosis. In the budding yeast Saccharomyces cerevisiae, endocytosis is localized to cortical actin patches, where a dense network of actin filaments is polymerized around clathrin-coated pits. This network is regulated by a complex set of proteins which cooperate in order to generate membrane invagination. We have recently developed in our laboratory a system in yeast extracts for the assembly of actin based structures around polystyrene microbeads. We believe that the protein composition of these structures could be similar to the protein composition found in vivo in cortical actin patches. We isolated the beads with their associated actin structures, and we would like to determine the protein composition by using mass spectroscopy. These results should allow us to further analyze the role of the different components during endocytosis.