Translation Initiation Factor IF3 Structure and Function

核糖体是一种复杂的分子机器，它忠实地将储存在遗传密码中的信息翻译成酶和其他蛋白质。 理解核糖体功能一直是并且仍然是分子生物学中最重要的挑战之一。 该项目的长期目标是了解核糖体如何确保其P位点的翻译准确性（正确解码）。 P位点在翻译起始期间解码信息，并且跨越二十多年的工作已经表明翻译起始因子3（IF 3）在这一事件中起着至关重要的作用，几乎可以肯定是通过调节核糖体本身固有的活动。 在过去的5年中，出现了重要的新见解，包括IF 3和核糖体的三维结构，IF 3/核糖体复合物的可视化，以及IF 3结构和功能的新见解。 IF 3被认为在P位点功能水平或接近P位点功能水平上起作用，最有可能改变核糖体构象，从而改变P位点。 这种P-位点活性的调节可能表现在几种看似无关的IF 3效应中。 这些发展为最终理解P-位点功能及其IF 3的调节提供了很大的希望。 然而，在合理地实现这些重要目标之前，必须首先解决第三次综合框架结构和功能的几个基本和关键方面。 该项目涉及对这一水平上几个主要未回答问题的综合遗传和生物化学攻击。 关键IF 3突变的影响将通过一系列强大的检测来检查，这些检测旨在监测IF 3在几个不同核糖体功能水平上的功能。 这些努力有助于测试IF 3对核糖体功能的调节的当前模型。 IF 3与核糖体相互作用所需的决定因素也将进行探索，并在核糖体本身的IF 3结合决定因素将被确定，在现有的实验证据的基础上的战略。 这些研究对于最终了解IF 3的结构和功能如何与核糖体整合至关重要。 特别是，这项工作将为最终直接在P位点解决IF 3功能的研究奠定坚实的基础。

AbstractMCB-0079305SimonsSIMONSThe ribosome is a complex molecular machine that faithfully translates information stored in the genetic code, into enzymes and other proteins. Understanding ribosome function has been and remains one of the most important challenges in molecular biology. The long-term goal of this project is to understand how the ribosome ensures translational accuracy (proper decoding) at its P-site. The P-site decodes information during translation initiation, and work spanning more that two decades has shown that translation initiation factor three (IF3) plays a crucial role in this event, almost certainly by modulating activities that are intrinsic to the ribosome itself. During the past 5 years, important new insights have emerged, including the three-dimensional structure of IF3 and the ribosome, visualization of the IF3/ribosome complex, and new insights into IF3 structure and function. IF3 is thought to operate at or close to the level of P-site function, most likely altering ribosome conformation and thereby the P-site. This modulation of P-site activity probably manifests itself in several seemingly unrelated IF3 effects. These developments hold great promise for an eventual understanding of P-site function and its modulation by IF3. However, before those important goals can be reasonably tackled, several basic and key aspects of IF3 structure and function must first be resolved. This project involves an integrated genetic and biochemical attack on several major unanswered questions at this level. The effects of key IF3 mutations will be examined with a powerful battery of assays designed to monitor IF3 function at several distinct levels of ribosome function. These efforts are instrumental in testing current models for IF3 modulation of ribosome function. IF3 determinants required for interaction with the ribosome will also be explored, and the IF3 binding determinants within the ribosome itself will be identified, in strategies based on existing experimental evidence. These studies are essential for an eventual understanding of how IF3 structure and function are integrated with that of the ribosome. In particular, this work will form a strong underpinning for studies that will eventually tackle IF3 function directly at the P-site.