Artie McFerrin Department of Chemical Engineering, Texas A&M University, College Station, TX, United States.
McKetta Department of Chemical Engineering, The University of Texas, Austin, TX, United States.
Methods. 2018 Jul 1;143:34-47. doi: 10.1016/j.ymeth.2018.01.015. Epub 2018 Feb 1.
There are over 150 currently known, highly diverse chemically modified RNAs, which are dynamic, reversible, and can modulate RNA-protein interactions. Yet, little is known about the wealth of such interactions. This can be attributed to the lack of tools that allow the rapid study of all the potential RNA modifications that might mediate RNA-protein interactions. As a promising step toward this direction, here we present a computational protocol for the characterization of interactions between proteins and RNA containing post-transcriptional modifications. Given an RNA-protein complex structure, potential RNA modified ribonucleoside positions, and molecular mechanics parameters for capturing energetics of RNA modifications, our protocol operates in two stages. In the first stage, a decision-making tool, comprising short simulations and interaction energy calculations, performs a fast and efficient search in a high-throughput fashion, through a list of different types of RNA modifications categorized into trees according to their structural and physicochemical properties, and selects a subset of RNA modifications prone to interact with the target protein. In the second stage, RNA modifications that are selected as recognized by the protein are examined in-detail using all-atom simulations and free energy calculations. We implement and experimentally validate this protocol in a test case involving the study of RNA modifications in complex with Escherichia coli (E. coli) protein Polynucleotide Phosphorylase (PNPase), depicting the favorable interaction between 8-oxo-7,8-dihydroguanosine (8-oxoG) RNA modification and PNPase. Further advancement of the protocol can broaden our understanding of protein interactions with all known RNA modifications in several systems.
目前已知的高度多样化的化学修饰 RNA 超过 150 种，它们具有动态性、可逆性，并能调节 RNA-蛋白质相互作用。然而，人们对如此丰富的相互作用知之甚少。这可以归因于缺乏允许快速研究可能介导 RNA-蛋白质相互作用的所有潜在 RNA 修饰的工具。作为朝着这个方向迈出的有希望的一步，这里我们提出了一种用于表征含有转录后修饰的 RNA 与蛋白质之间相互作用的计算方案。给定 RNA-蛋白质复合物结构、潜在的 RNA 修饰核糖核苷位置以及用于捕获 RNA 修饰能量学的分子力学参数，我们的方案分两个阶段运行。在第一阶段，一个决策工具，包括短模拟和相互作用能计算，以高通量的方式快速有效地搜索，通过根据其结构和物理化学性质分类成树的不同类型的 RNA 修饰列表，并选择一组易于与目标蛋白相互作用的 RNA 修饰。在第二阶段，使用全原子模拟和自由能计算详细检查被蛋白质识别为修饰的 RNA。我们在涉及与大肠杆菌（E. coli）蛋白 Polynucleotide Phosphorylase（PNPase）复合的 RNA 修饰研究的测试用例中实施并实验验证了该方案，描绘了 8-氧代-7,8-二氢鸟苷（8-oxoG）RNA 修饰与 PNPase 之间有利的相互作用。该方案的进一步发展可以拓宽我们对几个系统中所有已知 RNA 修饰与蛋白质相互作用的理解。
