甘草3-羟基-3-甲基戊二酰辅酶A还原酶基因多态性对其编码酶催化效率的影响

Researches on the influence of 3-hydroxy-3-methylglutary-coenzyme A reductase gene polymorphism on catalytic efficiency of its encode enzyme in Glycyrrhiza uralensis

目的:利用GC-MS方法分析甘草不同3-羟基-3-甲基戊二酰辅酶A还原酶(HMGR)基因型表达蛋白的催化效率,为揭示甘草HMGR多态性在优质甘草药材形成中的作用奠定基础。方法:以从甘草中克隆的4种HMGR基因突变型构建表达载体,转化Escherichia coli BL21,进行诱导表达、检测、纯化及体外酶促反应,采用TLC及GC-MS对反应产物进行定性及定量分析。结果:L/V型突变(-HSL,-HSV)催化活性近似,GA插入型突变(GALLV,GALSV)催化活性近似,但插入型突变的催化活性显著高于前者,是前者的2倍左右。结论:甘草功能基因HMGR基因多态性可能是甘草优质药材形成的分子基础。

Objective： To analyse the effect of expression proteins containing different escherichia coli of 3-hydroxy-3-methylglutary-coenzyme A reductase （HMGR） genic mutation on the conversion efficiency of MVA with GC-MS method,in order to lay a foundation for revealing the function of HMGR gene polymorphism of Glycyrrhiza uralensis in the production of high-quality G. uralensis medicines. Method： The expression carrier was established from four HMGR genic mutation types cloned from G. uralensis and transformed into Escherichia coli BI22l. The protein was induced to express, detected and purified. The purified protein was adopted for in vitro enzymatic reaction. TLC and GC-MS were used for qualitative and quantitative analysis on reaction products. Result： The catalytic activi- ty of L/V genotype（-HSL and -HSV） was similar, and so was the catalytic activity of the genotype with GA insertion（ GALLV and GALSV）, but the catalytic activity of the latter was around 2 times higher than that of the former. Conclusion： The functional gene poIymorphism of G. uralensis may be the molecular foundation for the production of high-quality G. uralensi medicines.