Cryo-EM analysis of ribosomal frameshifting

核糖体移码的冷冻电镜分析

基本信息

批准号：
BB/D009499/1
负责人：
Ian Brierley
金额：
$ 29.96万
依托单位：
University of Cambridge
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2006
资助国家：
英国
起止时间：
2006 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FD009499%2F1
关键词：
Cryo EM analysis ribosomal frameshifting

项目摘要

Cell proteins are encoded in DNA, the cell's genetic material, and expressed from so-called messenger RNA copied from DNA. This process of expression involves reading a triplet nucleotide code and translating it into an amino acid polymer, the protein. The process of translation involves structures called ribosomes. These move along the messenger RNA decoding the triplets and adding one amino acid for each triplet to the growing amino acid chain. Viruses that infect cells carry their own genetic material and occasionally, they make messenger RNAs that contain a specific signal (called a frameshift signal) that tells the ribosome to stop making one type of triplet and to start making another. The result is that one messenger RNA can make two proteins. This is typical of viruses; as their own genetic material is usually relatively small, they use all sorts of tricks to maximise the number of proteins they can make. In this project, we wish to examine what happens to the ribosome when it encounters a frameshift signal. The question is 'what happens to the ribosome that makes it misbehave?' To answer this, we will purify ribosomes caught in the act of translating through the frameshift signal and study them by microscopy. The microscopy technique we will use is based on electrons rather than light, so is extremely powerful, and is called cryo-electron microscopy. It is 'cryo' because the samples are frozen in aqueous solution before examination to keep them in a natural and stable state. The images we obtain will hopefully tell us something about how the frameshift signal interferes with the ribosome, and should also be informative about how ribosomes work during normal protein synthesis.

细胞蛋白编码在细胞的遗传材料DNA中，并根据从DNA复制的所谓信使RNA表示。这种表达过程涉及阅读三重核苷酸代码，并将其转化为氨基酸聚合物，即蛋白质。翻译过程涉及称为核糖体的结构。这些沿信使RNA移动，将三胞胎解码并为每个三重态添加一个氨基酸到生长的氨基酸链中。感染细胞携带自己的遗传物质的病毒，偶尔会产生信使RNA，其中包含特定信号（称为移码信号），该信号告诉核糖体停止制作一种类型的三重态并开始制作另一种三重态。结果是一个使者RNA可以生产两种蛋白质。这是病毒的典型特征；由于他们自己的遗传物质通常相对较小，因此他们使用各种技巧来最大程度地提高其可以制造的蛋白质数量。在这个项目中，我们希望检查核糖体遇到移码信号时会发生什么。问题是“使其表现不佳的核糖体会发生什么？”为了回答这一点，我们将净化核糖体在翻译信号的行为中捕获，并通过显微镜研究它们。我们将使用的显微镜技术是基于电子而不是光的，因此非常强大，被称为冷冻电子显微镜。这是“冷冻”，因为在检查之前将样品冷冻在水溶液中，以使它们保持自然而稳定。我们获得的图像有望告诉我们一些有关框架信号如何干扰核糖体的信息，并且还应该了解核糖体在正常蛋白质合成过程中的工作方式。