Dissecting the Molecular Mechanism of Protein-DNA Interactions

基因表达调控的一个重要组成部分是蛋白质-DNA相互作用。Cys2-His2锌指转录因子(ZF)家族是真核生物中最大的核酸结合蛋白家族，参与了大多数基因的调控，在多种刺激下被激活。从结构的角度来看，这个转录因子家族是高度保守的，由两个或更多ZF模块结构域组成，它们共同识别特定的DNA序列。显然，ZFS是研究非特异性和特异性蛋白质-DNA结合的基本原理的理想模型系统，并揭示了基因转录调控的关键步骤。该项目的目标是利用基于结构的计算方法，结合来自生化实验的现有选择和结合数据，以及蛋白质数据库中存储的高分辨率晶体结构，揭示负责蛋白质识别和结合的热力学和动力学的微调分子相互作用。具体地说，将使用分子动力学采样作为指导原则来研究反离子和侧链在蛋白质-DNA结合中的作用，以评估和预测由于点突变而导致的结合自由能的变化，使我们能够更详细地探索它们相互作用的分子机制。为了了解蛋白质如何调控几乎所有的生物过程，了解蛋白质与DNA的相互作用至关重要。到目前为止，这个问题已经通过生化实验、具有挑战性和昂贵的络合物结构确定来研究，而计算研究主要集中在基于序列的基序算法上。这个项目将结合动力学和分子生物物理学，以揭示可能逃脱当前方法的序列/结构关系。在这一根本问题上的任何进展都必然会带来对基因调控如何在细胞中协同工作的更好理解。该项目将涉及对研究生进行计算/结构建模方面的培训，并将努力从该项目中代表性不足的群体中招募有才华的学生。该项目还可以在开发新的药物方面产生广泛的影响，这些药物可以阻断或下调与致病途径或蛋白质相关的特定DNA序列的表达。

A critical component on the regulation of gene expression is protein-DNA interactions. The Cys2-His2 zinc finger (ZF) family of transcription factors is the largest family of nucleic acid binding proteins in eukaryotes and a key participant in the regulation of most genes, being activated in response to a wide variety of stimuli. From a structural point of view, this family of transcription factors (TFs) is highly conserved, consisting in two or more ZF modular domains that work together to recognize specific DNA sequences. It is then apparent that ZFs are an ideal model system to study the fundamental principles governing non-specific and specific protein-DNA binding, and shed light into an essential step in the transcription regulation of genes. The goal of this project is to reveal the finely tuned molecular interactions responsible for the thermodynamics and dynamics of protein recognition and binding using structural-based computational approaches in combination with available selection and binding data from biochemical experiments, and high resolution crystal structures deposited in the Protein Data Bank. Specifically, the role of counter ions and side chains in protein-DNA association will be studied using molecular dynamics sampling as a guiding principle to assess and predict changes in binding free energy due to point mutations, allowing us to explore in much greater detail the molecular mechanism of their interaction. Understanding protein-DNA interactions is crucial in order to learn how proteins regulate almost every biological process. So far this problem has been studied by biochemical experiments, challenging and expensive structural determination of the complexes, and computational studies focused mostly in sequence-based motif algorithms. This project will incorporate dynamics and molecular biophysics to uncover sequence/structural relationships that might escape current methods. Any progress in this fundamental problem is bound to bring about a better understanding of how gene regulation works cooperatively in a cell. The project will involve the training of graduate students in computational/structural modeling, and efforts will be made to recruit talented students from underrepresented groups in the project. This project can also make a broad impact in the development of new agents that block or down regulate the expression of specific DNA sequences linked to a pathogenic pathway or protein.