Interactions Regulating Translation and Protein Biogenesis in Vivo

调节体内翻译和蛋白质生物发生的相互作用

基本信息

批准号：
8489518
负责人：
Allen Rowdon Buskirk
金额：
$ 80.06万
依托单位：
UNIVERSITY OF NOTRE DAME
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-06-10 至 2018-04-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Proper cellular function requires efficient folding of cellular proteins. In the cell, protein foldng starts cotranslationally, concomitantly with protein synthesis by the ribosome, following pathways that can be distinct from the refolding of full-length proteins in vitro. The average, bul translation rate in E. coli is -20 aa/sec, but this rate can vary by more than an order of magnitude for the translation of specific mRNA sequences. Recent results have highlighted that altering the translation rate of small portions of some genes can significantly affect the folding efficiency of the encoded protein (correct folding versus misfolding and aggregation, or degradation), or alter the partitioning between two alternative folded structures. Yet despite the potential impact of local translation rate on protein biogenesis in vivo, the interactions between the ribosome and mRNA and/or nascent chain sequences that control local translation rate remain opaque. Nor do we know what fraction of proteins in the proteome has co-translational folding mechanisms that are significantly affected by altered translation rate. The Pis of this proposal have constructed a network to span their established expertise in genetics, molecular biology, genomics, biochemistry, biophysics, and organic, analytical and physical chemistry, creating a team uniquely suited to tackle three significant outstanding questions regarding the mechanisms and outcomes of altered translation rate in E. coli. Our network will determine: (1) What specific features of mRNA and/or nascent chain sequences shape absolute local translation rate in vivo, and by what mechanisms? (2) What proteins in the proteome are most susceptible to aggregation or degradation when translation rate is altered? (3) To what extent does translational pausing alter the conformation of the ribosome and/or its interactions with other proteins in vivo? Taken together, results from this proposal will represent crucial steps towards a comprehensive picture of translation rate control in E. coli, and the effects of altered translation rate on protein biogenesis, including the heterologous expression of human proteins. PUBLIC HEALTH RELEVENCE: The rate of protein synthesis is not uniform, but we do not fully understand why some sequences are translated more slowly than others. Altering local translation rate can affect protein folding, and hence cell function. We will develop new approaches to measure local translation rates and the mechanisms that determine these rates, and identify proteins whose folding is most affected by translation rate differences.

描述（由申请人提供）：正确的细胞功能需要有效折叠细胞蛋白。在细胞中，蛋白质折叠从核糖体的蛋白质合成开始，沿核糖体的蛋白质合成，遵循可以与全长蛋白重新折叠的途径。大肠杆菌中的平均体积平移速率为-20 aa/sec，但对于特定mRNA序列的翻译，该速率的变化大于一个数量级。最近的结果表明，改变某些基因的小部分的翻译速率可以显着影响编码蛋白的折叠效率（正确的折叠与错误折叠和聚集或降解），或更改两个替代折叠结构之间的分配。尽管局部翻译速率对体内蛋白质生物发生有潜在的影响，但控制局部翻译速率的核糖体和mRNA和/或新生链序列之间的相互作用仍然不透明。我们也不知道蛋白质组中哪些蛋白质的比例具有共同翻译机制，这些机制显着受翻译速率的影响。该提案的PI构建了一个网络，以涵盖其在遗传学，分子生物学，基因组学，生物化学，生物物理学以及有机，分析和物理化学方面的既定专业知识，从而创建了一个独特的团队，该团队适合解决大肠杆菌中有关机制的机制和影响率变化的三个重要问题。我们的网络将确定：（1）mRNA和/或新生链序列在体内形成绝对局部翻译率的哪些特定特征，以及通过什么机制？（2）当翻译速率改变时，蛋白质组中哪些蛋白最容易聚集或降解？（3）翻译暂停的程度在多大程度上改变了核糖体和/或其与其他蛋白质的相互作用的构象？综上所述，该提案的结果将代表迈向大肠杆菌中翻译速率控制的全面图片的关键步骤，以及转化速率改变对蛋白质生物发生的影响，包括人类蛋白质的异源表达。公共卫生相关性：蛋白质合成的速率不是均匀的，但是我们不完全理解为什么某些序列比其他序列更慢。改变局部翻译率会影响蛋白质折叠，从而影响细胞功能。我们将开发新的方法来衡量局部翻译速率和确定这些速率的机制，并确定其折叠率最大的蛋白质受翻译率差异的影响。