Structure-based Simulation of Riboswitches: Electrostatic Effects

随着RNA生物学、RNA生物技术和RNA生物医学的发展， 我们迫切需要尽可能详细地了解RNA的作用机制。小RNA 技术（miRNAs和siRNAs）和CRISPR-Cas9基因编辑正在改变人类基因组的格局。 生物技术和生物医学。核糖开关RNA（i）在细菌基因调控中很重要，（ii） 感兴趣的抗微生物靶点，以及（iii）具有优化生物技术的潜力， CRISPR-Cas9.这些RNA是研究RNA特征的极好模型系统 机制：镁驱动的静电效应，大的3-D构象变化， 二级结构和共转录效应。自2009年以来，我们发布了各种 显式溶剂分子动力学模拟、基于结构的分子模拟和湿实验室 核糖开关的生物化学研究，以了解它们的运作和镁对 核糖开关的结构和功能。主要关注SAM-1核糖开关的适体结构域， 我们已经确定，镁和SAM一起工作，以实现完全崩溃，原生 状态此外，镁促进部分塌陷，使适体处于允许的状态 通过表达平台进行链侵入。在这个项目中，我们将研究整个核糖开关 （适体和表达平台），研究镁在核糖开关功能中的作用。使用 基于原子结构的RNA和镁的静电势模型，我们将解开 内球、外球和扩散镁离子效应在核糖开关操作中的作用。 利用实验确定的中间构型，我们将研究 在核糖开关功能的各个点的中间体。我们的建模，我们将增强 各种生物化学和生物物理实验的限制。我们将解决 一个基本的问题是，离子环境如何使核糖开关RNA完成它们的功能？ 功能？

With new developments in RNA biology, RNA biotechnology and RNA biomedicine each year, there is an urgent need to understand RNA mechanism in as much detail as possible. Small RNA techniques (miRNAs and siRNAs), and CRISPR-Cas9 gene editing are changing the landscape of biotechnology and biomedicine. Riboswitch RNAs are (i) important in bacterial gene regulation, (ii) interesting antimicrobial targets, and (iii) have potential for optimizing biotechnologies such as CRISPR-Cas9. These RNAs are excellent model systems for studying the hallmarks of RNA mechanism: magnesium-driven electrostatic effects, large 3-D conformational changes, changes in secondary structure, and co-transcriptional effects. Since 2009, we have published a variety of explicit solvent molecular dynamics simulation, structure-based molecular simulation, and wetlab biochemistry studies of riboswitches to understand their operation and the effect of magnesium on riboswitch structure and function. Focusing mainly on the aptamer domain of the SAM-I riboswitch, we have established that magnesium and SAM work together to achieve the fully collapsed, native state. In addition, magnesium facilitates a partial collapse, leaving the aptamer in a state permissible to strand invasion by the expression platform. In this project, we will study the entire riboswitch (aptamer and expression platform), investigating the role of magnesium in riboswitch function. Using atomistic structure-based electrostatic potential models for RNA and magnesium, we will disentangle the roles of inner sphere, outer sphere and diffuse magnesium ion effects in riboswitch operation. Using experimentally determined intermediate configurations, we will study transitions between intermediates during various points of riboswitch function. Our modeling we be enhanced by constraints from a variety of biochemical and biophysical experiments. We will address the fundamental question, how does the ionic environment enable riboswitch RNAs to accomplish their function?