细胞粘附蛋白解折叠的单分子力谱研究

项目名称： 细胞粘附蛋白解折叠的单分子力谱研究

项目编号： No.21204034

项目类型： 青年科学基金项目

立项/批准年度： 2013

项目学科： 高分子科学

项目作者： 刘宁宁

作者单位： 辽宁石油化工大学

项目金额： 25万元

中文摘要： 利用特殊的细胞表面分子产生细胞粘附作用在生命科学领域中是具有重要地位的。致病菌白色念珠菌依靠其表面的一种Als蛋白与宿主细胞进行粘附，而粘附作用主要依靠蛋白中的TR区域。基于原子力显微镜的单分子力谱技术是一种有效地在单分子水平上研究分子间/分子内相互作用的方法。本项目中，我们首先欲利用单分子力谱技术对活体细菌表面的TR区域进行往复拉伸，从而探测解折叠的蛋白是否可以重新折叠回去。若该过程可逆，欲利用单分子力谱中的力钳模式记录TR区域解折叠-折叠随时间变化的轨迹，并探测折叠过程中所用外力对折叠停留时间的影响。蛋白的解折叠过程是具有拉力依赖性的，欲通过在不同的恒定力值下对TR区域进行拉伸，从而测得解折叠概率随时间变化的函数以及每个恒力下的速率常数,并结合Arrhenius方程计算出解折叠的动力学参数并绘制其能垒图。

中文关键词： 烟草花叶病毒；单分子力谱；解组装；动力学；力钳模式

英文摘要： Cell adhesion mediated by speci?c cell surface molecules plays a pivotal role in life sciences. Cellular communication, tissue development, in?ammation, cancer and microbial infection are just a few examples of cellular processes regulated by cell adhesion molecules. Adhesion of C. albicans yeast cells to host tissues is a key factor in the maintenance of commensal and pathogenic states, which is mediated by a family of cell surface proteins known as the agglutinin-like sequence (Als) proteins. Of the Als proteins, the cell-cell aggregation is mainly participated by a tandem repeat (TR) region. Atomic force microscopy (AFM)-based single molecule force spectroscopy (SMFS) is a very effective technique for the investigation of inter- or intramolecular interactions at sinlge molecule level. In this project, we firstly want to stretch and relax the TR region of Als protein on the surface of living cell repeatly to detect whether the unfolding structure can go back to its folding state. If this unfolding-folding process is reversible, we want to use the force-clamp mode of SMFS techniques to record the unfolding-folding trajectory of TR region as a function of time, and to investigate the foce-dependence of duration for folding collapse. For the mechanical unfolding is a force-dependent process, we plan to stretch th

英文关键词： Tobacco mosaic virus；single molecule force spectroscopy；disassembly processes；dynamics；force-clamp