Disassembly and Degradation of Photosystem I

描述（由申请人提供）：该项目的长期目标是了解膜蛋白复合物是如何分解和降解的。许多蛋白质以一种受调节和消耗能量的方式降解，细胞识别为异常的蛋白质被这些“质量控制”系统优先靶向。尽管用于降解可溶性蛋白的细胞机制已经被很好地理解，但对膜蛋白相应过程的了解却滞后，尽管许多膜蛋白在信号转导、生物能量学和细胞稳态等重要过程中发挥着关键作用。一些人类疾病，如囊性纤维化和视网膜色素变性，是由膜蛋白的点突变降解引起的。本文提出的模式系统是单细胞绿藻衣藻（Chlamydomonas reinhardtii）中的光系统I （PS1）复合体。这种蛋白质是光合作用电子传递链的重要组成部分，它利用与其相关的叶绿素分子吸收的光来驱动电子在类囊体膜上的转移。它应该作为膜蛋白降解的一个很好的模型底物，因为蛋白质的功能部分在膜的平面上，在那里它有许多光谱探针。该项目利用了抑制基因和生物化学的互补方法。该项目的具体目标是：(1)完成一系列抑制因子突变的遗传表征，这些突变显示PS1的降解效率降低；(2)使用基于分子图谱的技术克隆具有代表性的抑制因子突变；(3)创建生化互补系统，并将其用于纯化涉及靶向和降解PS1的蛋白质。

DESCRIPTION (provided by applicant): The long-term goal of this project is to understand how membrane protein complexes are disassembled and degraded. Many proteins are degraded in a regulated and energy-consuming fashion, and proteins recognized by the cell as aberrant are targeted by these "quality control" systems preferentially. Although the cellular mechanisms used to degrade soluble proteins are becoming well understood, knowledge of the corresponding process for membrane proteins has lagged, despite the key role many membrane proteins play in important processes, such as signal transduction, bioenergetics, and cellular homeostasis. Several human diseases, such as cystic fibrosis and retinitis pigmentosa, are caused by degradation of membrane proteins with point mutations. The model system proposed here is the Photosystem I (PS1) complex in the unicellular green alga, Chlamydomonas reinhardtii. This protein is an essential part of the photosynthetic electron transport chain and uses light absorbed by its associated chlorophyll molecules to drive electron transfer across the thylakoid membrane. It should serve as an excellent model substrate for membrane protein degradation, because the functional portion of the protein is in the plane of the membrane, where it has many spectroscopic probes built into it. The project makes use of the complementary approaches of suppressor genetics and biochemistry. The specific aims of the proposed continuation project are (1) to finish genetic characterization of a set of suppressor mutations that display decreased efficiency of degradation of PS1, (2) to clone a representative suppressor mutation using molecular map-based techniques, and (3) to create a biochemical complementation system and use it for the purification of protein(s) involved in targeting and degrading PS1.