Mechanism and Structural Dynamics of Translation

理解和描述大分子机器设计背后的动态特性是当前生物物理研究和生物分子工程的前沿。这项拟议中的研究侧重于细胞组件，这些组件是细胞机制的一部分，用于解码遗传信息，并在称为翻译的过程中合成功能蛋白质。翻译是核糖体的一部分，核糖体是所有活细胞必不可少的大分子复合体。核糖体由蛋白质和RNA组成，这是两类主要的功能生物分子。结构研究表明，核糖体是一个高度动态的生物分子装配线，其中的两个关键功能(肽键的形成和解码)似乎完全由RNA来执行。然而，为了在活细胞中观察到快速而准确的翻译，需要额外的蛋白质因素。起始因子(IF)和释放因子(RF)参与蛋白质合成的起始和终止，而伸长因子(EF)则是肽链伸长的循环过程所必需的。这些翻译因子中有许多是核苷酸水解酶，起着分子开关的作用，对特定的输入信号和特定的输出做出反应。这些信息处理和决策事件的动态对于我们理解翻译和生物分子机器性能背后的设计原理非常有意义。我们刚刚开始了解在分子水平上传递信息的高度动态网络的设计是如何涉及到翻译等分子过程的。因此，拟议的研究将针对这些通信网络，并提供使其能够进行详细分析的分子工具。*拟议的研究计划通过研究细菌EF-Tu和结构相关的翻译GTP酶的结构动力学和生化特性来应对这一挑战。翻译GTP酶构成了一类不同的调节酶，尽管它们具有共同的结构设计，但在核糖体依赖的蛋白质合成过程中具有非常不同的酶性质和功能。为了确定特定的酶特性和机制作用如何植根于特定的翻译GTP酶的动态特性以及潜在的设计策略，拟议的研究计划将结合最先进的生物物理技术，如荧光光谱、单分子荧光共振能量转移(sm-fret)、快速动力学和分子动力学(MD)模拟。*了解核糖体等大分子组装过程的动力学特性对于蛋白质/RNA折叠和功能以及新型生物分子纳米机器的合理设计具有重要意义。**

Understanding and describing the dynamic properties underlying the design of macromolecular machines is a current frontier in biophysical research and biomolecular engineering. The proposed research focuses on the cellular components that are part of the cells machinery to decode genetic information and to synthesize functional proteins during a process called translation. Translation takes part at the ribosome, a large macromolecular complex that is essential to all living cells. The ribosome is composed of proteins and RNAs, the two major classes of functional biomolecules. Structural studies have revealed that the ribosome is a highly dynamic biomolecular assembly line in which the two key functions (peptide-bond formation and decoding) seem to be carried out solely by RNA. However, for the rapid and accurate translation observed in living cells, additional protein factors are required. Initiation (IF) and release factors (RF) are involved in initiation and termination of protein synthesis, whereas elongation factors (EF) are required for the cyclic process of peptide chain elongation. Many of these translation factors are nucleotide hydrolases that function as molecular switches, responding to a particular input signal with a specific output. The dynamics of these information processing and decision-making events is of great interest for our understanding of translation and the design principles underlying the performance of biomolecular machines. We are just beginning to understand how the design of highly dynamic networks relaying information at the molecular scale is involved in molecular process such as translation. The proposed research will therefore target these communication networks and provide molecular tools enabling their detailed analysis.***The proposed research program addresses this challenge by studying the structural dynamics and biochemical properties of bacterial EF-Tu and structurally related translational GTPases. Translational GTPases constitute a diverse class of regulatory enzymes that although sharing a common structural design, have vastly different enzymatic properties and functions during ribosome-depend protein synthesis. In order to identify how the particular enzymatic properties and mechanistic roles are rooted in the dynamic properties of the particular translational GTPase and what the underlying design strategies are, the proposed research program will use a combination of state-of-the-art biophysical techniques such as fluorescence spectroscopy, single molecule Fluorescence Resonance Energy Transfer (sm-FRET), rapid kinetics and molecular dynamics (MD) simulations. ***Understanding the dynamical properties of processes in macromolecular assemblies like the ribosome will be of great importance for protein / RNA folding and function as well as rational engineering of novel biomolecular nano-machines.**