BLM解旋酶对G4 DNA解旋的分子作用机理研究

解旋酶属于生物马达蛋白，它们利用水解NTP产生的能量打开核酸双链或其它核酸结构，在生物体内参与核酸复制、转录、修复、重组、核糖体组装、蛋白质翻译、端粒稳定等很多过程。本项目所研究的BLM解旋酶是人类细胞中与DNA复制、修复和重组有关的重要蛋白。与其重要功能相对应，BLM是少有的几种能够识别和解开多种特异性DNA底物结构的人类解旋酶之一。在BLM所有的DNA底物中，G-四联体（G4）与双链差别最大、结构也最复杂，有关BLM识别和解开G4结构的分子机理也成为解旋酶领域中备受关注的问题之一。在本申请项目中，我们将利用单分子荧光、磁镊、快速停流反应动力学以及动态光散射等方法，深入研究BLM与G4的结合特性、对G4的解旋过程以及它的构象变化，由此揭示BLM识别、解旋G4的分子作用机制，为更好地理解BLM蛋白结构与功能的关系提供帮助。

Helicases are molecular motors. They disrupt nucleic-acid duplexes and other structures using the energy derived from NTP hydrolysis. They are involved in many life processes such as nucleic-acid replication, transcription, repair, recombination, ribosome biogenesis, protein translation, telomere stabilization. The BLM helicase to be studied in the present project is an important protein involved in DNA replication, repair and recombination in human cells. In accordance with its important functions, BLM is one of the few human helicases that are able to recognize and unwind many types of specific DNA substrates. Among all the substrates for BLM, G-quadruplex (G4) is the most different from duplex and has the most complex structure. The molecular mechanism for recognization and unwinding of G4 by BLM is becoming one of the topics of most concern in the helicase research field. In the present project, we will use single-molecule fluorescence, magnetic tweezers, stopped-flow kinetic and dynamic light scattering methods to study intensively the G4 binding properties, the G4 unwinding processes, as well as conformational changes of BLM, so as to reveal the molecular mechanism for recognization and unwinding of G4 DNA by BLM and provide clues for better understanding the protein structure and function relationships for BLM.