人工核酶M_1GS结构与功能相互关系的研究  

Interrelation between the structure and the function of artificial ribozyme M_1GS

作　　者：李弘剑[1] 孙亮[1] 李月琴[1] 郑永霞[1] 唐冬生[1] 张欣[1] 周天鸿[1] 

机构地区：[1]暨南大学生命科学技术学院,广东广州510632

出　　处：《中国病理生理杂志》2006年第6期1049-1054,共6页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金项目资助(No.30370776);广东省自然科学基金重大项目资助(No.36703);广东省自然科学基金项目资助(No.021162;No.000718)

摘　　要：目的:研究核酶M1GS其二级结构与体外切割活性之间的关系。方法:以人巨细胞病毒(HCMV)DNA聚合酶基因UL54为靶基因,构建了人工核酶M1GS-T7。通过软件RNAstructure对M1GS在3个具有相对稳定结构的温度(20℃、37℃、55℃)的空间构象进行模拟,然后通过体外切割实验来检测不同温度下核酶M1GS体外切割活性的变化。为一步研究核酶M1GS二级结构与体外切割活性之间的关系,参照温度变化实验结果及RNA二级结构的模拟结果,引入突变位点,构建了在37℃与55℃时M1GS-T7具有相同二级结构的突变型核酶mM1GS-T7,并通过体外切割实验对两者的活性进行比较。结果:在温度变化实验中,55℃核酶的体外切割活性最高。而在突变实验中,37℃mM1GS-T7比M1GS-T7的活性略高。结论:具有某种特定二级结构的M1GS-T7有相对较高的体外切割活性,核酶的结构与其功能之间存在一定的对应关系。AIM： To study the interrelation between the structure and the function of artificial ribozyme M1GS. METHODS： Ribozyme Mi GS - T7, which targeted the mRNA segment of HCMV UL54 gene, was constructed. The secondary structure of MIGS was simulated under different temperatures （20 ℃, 37 ℃ or 55℃, at which the secondary structure of MIGS was relatively stable） and the interrelation between the secondary structure and the cleavage activity of MIGS was analyzed under different temperature /n vitro. To investigate the interrelation between the structure and the function of ribozyme Mi GS further, mMi GS - T7 was designed, in which some mutation sites was added, according to the result of temperature change experiment and the simulated secondary structure, showing that were the same structures at 37℃ as that of MIGS- T7 at 55℃. RESULTS： In temperature change experiment, the cleavage activity of MIGS- T7 was highest at 55℃. The result of mutant experiment showed that the mutant type was more active than M1GS - T7 at 37℃. CONCLUSION： The cleavage activity of MIGS, which has some certain secondary structure, is higher than others. There is some interrelation between the structure and the function of MIGS.

关 键 词：温度 突变 基因 U154 RNA 催化 

分 类 号：R363[医药卫生—病理学]