体外装配核小体过程组蛋白浓度依赖的动力学模型  

Dynamic Model of Histone-dependent Nucleosome Assembly in vitro

作　　者：张凤慧 郭明欣 赵宏宇 蔡禄 Zhang Fenghui;Guo Mingxin;Zhao Hongyu;Cai Lu(School of Life Science and Technology,Inner Mongolia University of Science and Technology,Baotou,014010)

机构地区：[1]内蒙古科技大学生命科学与技术学院

出　　处：《基因组学与应用生物学》2019年第7期2996-3001,共6页Genomics and Applied Biology

基　　金：国家自然科学基金项目(31760247; 61671256; 61662055);内蒙古自然科学基金项目(2017MS0303)共同资助

摘　　要：为探索组蛋白浓度对核小体体外装配的影响,本研究表达纯化了4种组蛋白,通过控制实验反应体系中组蛋白的浓度,利用盐透析法在体外装配了核小体,检测分析了组蛋白浓度与核小体组装效率的关系。以此实验数据为基础,提出了核小体组装过程组蛋白浓度依赖性的动力学模型。实验结果发现,反应体系中组蛋白浓度与核小体生成量呈典型的线性关系。依据动力学理论模型,进行线性回归拟合,回归系数达到0.963;经计算601 DNA序列组装核小体的反应速率常数k为1.49×10^-5mL·h·μg^-1。CS1序列验证动力学模型的线性回归相关系数为0.989,反应速率常数为1.52×10^-5mL·h·μg^-1。该实验方法及动力学模型中反应速率常数k可用于评价相同长度的DNA序列组装核小体的能力、组蛋白与其突变体以及组蛋白变体之间形成核小体结构能力的差异。该动力学模型的建立为理解核小体装配、核小体定位、染色质结构等相关问题提供了理论指导。Nucleosome is the fundamental repeating unit of eukaryot chromatin.By changing the access of trans-acting factors to their template and controlling the degree of chromatin opening or folding,nucleosome assembly regulates many biology processes occurred in chromatin level,such as DNA replication,RNA transcription,DNA repair and DNA recombination.Histones are the basic components in nucleosome,and investigating the effect of histone concertration on nucleosome assembly is the basis for understanding the dymanic of nucleosome assembly.Four histones were expressed and purified from Escherichia coli BL21 cells containing pET-histone expression plasmids.The nucleosomes were assembled with different histone concentration using the salt-dialysis method in vitro.Based on the quantificational results of efficiency of the nucleosome assembly and hisone concentration,a dynamical model of histone-dependent nucleosome assembly was proposed.The experimental results found that the ralation between efficiency of the nucleosome assembly and histone concentration showed typical linearity,and the regression coefficient was 0.963 in the equation of linear regression.The reaction rate constant k of nucleosome assembly on 601 DNA template was 1.49×10^-5mL·h·μg^-1.Verification test on CS1 DNA sequence showed that the regression coefficient of equation of linear regression was 0.989 and the reaction rate constant k on CS1 DNA template was 1.52×10^-5mL·h·μg^-1.The reaction rate constant k in dynamical equation can be employed to evaluate the ability of DNA sequences to assemble nucleosome,efficency of nucleosome formation on between histone and histone mutant or histone variants.The dynamical model may be helpful for understanding the regulation mechanism of nucleosome assembly,nucleosome positioning and chromatin structure.

关 键 词：组蛋白 核小体装配 染色质结构 

分 类 号：Q343.2[生物学—遗传学]