RNA聚合酶转录非平衡态物理机制解析

Gene transcription is one of the most fundamental and essential processes in molecular genetics and evolution, conducted by molecular motors in charge of the processes, i.e., mainly RNA polymerase, that work at non-equilibrium steady state. Since a nucleotide addition cycle occurs in several milliseconds and some intermediate states exists very shortly (less than 1microsecond), current experimental technologies can not observe these process directly. In recent years, rapid advancements in experimental technology at single molecule level provide abundant structural and dynamics information for the polymerases, making it possible to study their function dynamics at molecular level from a physics point of view. The major issues of our concern include non-equilibrium dynamical properties of small systems, the physical mechanisms of efficient propagation of genetic information as well as molecular responses and coordination. Our research will start from utilizing single molecule experimental data, and build models by combing information from both stochastic dynamics and molecular structures. We will employ techniques such as molecular dynamics simulation from atomistic to some coarse-grained levels to explore the mechanism of gene transcription. Through mimic the actual transcription condition, we will investigate the gene transcription mechanism from free energetic and structural evolution, dissecting underlying mechanisms and building correspondence between the phenomenology and structural & dynamics details. The study is helpful to understand and interpret biological phenomena, providing a basis for interpretation of experimental results and settling the foundation for further studying gene replication and transcription.

基因转录是分子生物遗传和进化中最基本和核心的过程，主要由RNA聚合酶在非平衡稳态下工作完成。由于其机理复杂，尺寸小（分子量级），周期短（毫秒级），反应各阶段快慢不一，部分中间态存在时间甚至小于微秒级，无法通过现有实验技术观测。随着计算机硬件、模拟技术的飞速发展和生物分子力场的完善，使得从微观角度来研究分子功能、探讨微观机理成为可能。本项目拟从单分子实验数据入手，结合体系的随机动力学和分子结构信息进行建模。采用全原子分子动力学，马尔可夫模型等相结合方法对基因转录机理进行探讨。通过模拟真实环境下的转录过程，从自由能和结构演化角度研究基因转录机理，从微观尺度上探讨基因转录非平衡态动力学特性，信息流的有效传递以及分子结构感应和协调的物理机制，为唯象模型提供具体的表征。本课题的开展将为理解和解释生命现象、预测实验结果提供依据，为深入研究基因复制和转录打下良好理论基础。

基因转录是遗传信息从DNA传递到RNA的过程，是分子生物遗传和进化中最基本和核心的步骤。由于转录机理复杂，体系尺寸小，周期短，各阶段快慢不一，部分中间态存在时间甚至小于微秒级，不利于实验技术进行机制探讨。对于转录机理的仍缺乏连续动态的认识，许多转录细节亟待进一步研究。本项目从单分子实验数据入手，结合体系的随机动力学和分子结构信息进行建模，采用全原子分子动力学，伞形采样技术，MMPBSA技术，马尔可夫模型等相结合方法对基因转录机理进行探讨。首先研究了聚合酶对NTP作用机制，发现它们之间建立广泛的相互作用为聚合反应创造了稳定的环境；然后研究PPi释放机理，发现PPi释放是一个非自发的过程；对双镁离子分离机理进行了研究，发现两个镁离子位置的不对等，导致反应结束后一个镁离子随PPi释放，另外一个镁离子留着活性位点；对NTP分子插入机理研究，发现三磷酸根和活性位点的相互作用有助于帮助NTP进行调至正确构象；从自由能角度阐述了RNAP对正确和非正确底物的识别，发现正确的NTP和活性位点结合更紧密，发现Tyr639对NTP预检测的有利证据。综合上述结果，本研究为探讨从自由能和结构演化角度研究基因转录机理，从微观尺度上探讨其非平衡态动力学特性，信息流的有效传递以及分子结构感应和协调的物理机制，为唯象模型提供具体的表征。此外本课题将为理解和解释生命现象、预测实验结果提供依据，为新药开发提供靶点和思路，为深入研究基因复制和转录打下良好理论基础。项目资助发表SCI论文8篇，待发表两篇。培养硕士博士研究生4名，其中2名已经毕业并取得硕士学位，2名在读。开展了和中科院，川大等国内知名机构的科研合作，顺利完成预定目标。由于基因转录是一项大工程，我们的研究还在继续深入，希望得到明年面上项目的资助。