Cryo-EM analysis of ribosomal frameshifting

细胞蛋白质在细胞的遗传物质DNA中编码，并由从DNA复制的所谓信使RNA表达。这个表达过程包括阅读一个三联核苷酸密码，并将其翻译成氨基酸聚合物，即蛋白质。翻译的过程涉及一种叫做核糖体的结构。它们沿着信使RNA移动，解码三联体，并为每个三联体添加一个氨基酸到不断增长的氨基酸链中。感染细胞的病毒携带自己的遗传物质，偶尔，它们会产生含有特定信号（称为移码信号）的信使RNA，该信号告诉核糖体停止制造一种类型的三联体并开始制造另一种。结果是一个信使RNA可以产生两种蛋白质。这是典型的病毒;由于它们自身的遗传物质通常相对较小，它们使用各种技巧来最大限度地增加它们所能制造的蛋白质数量。在这个项目中，我们希望研究当核糖体遇到移码信号时会发生什么。问题是"核糖体发生了什么事，使它行为不端？“为了回答这个问题，我们将纯化通过移码信号翻译的核糖体，并通过显微镜进行研究。我们将使用的显微镜技术是基于电子而不是光，所以非常强大，被称为冷冻电子显微镜。之所以称之为“cryo”，是因为在检测前，样品被冷冻在水溶液中，以保持其自然和稳定的状态。我们获得的图像有望告诉我们移码信号如何干扰核糖体，也应该是关于核糖体在正常蛋白质合成过程中如何工作的信息。

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.