大麦β-淀粉酶基因在大肠杆菌中的异源表达

Heterologous expression of β-amylase gene from barley in E. coli

大麦来源的β-淀粉酶具有稳定性高和工业应用效果好的优点,被广泛应用于食品、酿造以及粮食加工,但是其制备成本高,价格较昂贵。该研究使用大肠杆菌异源表达大麦来源的β-淀粉酶。首先通过基因合成技术获得密码子优化的大麦来源的β-淀粉酶基因,将该基因通过质粒p ET28a(+)在大肠杆菌BL21(DE3)中过量表达获得重组β-淀粉酶。其次,对重组表达的β-淀粉酶和大麦中直接提取的β-淀粉酶进行酶学性质比较分析。结果表明,重组表达的β-淀粉酶其分子质量大小与大麦中β-淀粉酶一致,即59 k Da,重组的β-淀粉酶比酶活由大麦来源的588 U/mg降为285.5 U/mg,最适作用温度降低了10℃,最适p H保持一致。所以,大麦β-淀粉酶能够在细菌中高效表达,但是其酶学性质发生较大改变。

Barley β-amylase is widely applied in food and brewing industries due to its high activities,good stability and performance. However,the low production rate and high price of barley β-amylase limit its application. In this study,the barley β-amylase was heterologously expressed in Escherich coli. The β-amylase gene was codon-optimized,inserted into plasmid p ET28a（ ＋） and over-expressed in E. coli BL21（ DE3） cells. After comparing the recombinant β-amylase with the native one from barley,the results showed that the molecular weight of the recombinantβ-amylase was similar with that of the native barley β-amylase,which was about 59 k Da. Its specific activity was decreased to 285. 5U / mg,which was lower than that of the enzyme from barley（ 588 U / mg）. Furthermore,its optimum temperature was declined 10 ℃,while its optimum p H kept consistent. Therefore,barley β-amylase gene could be effectively over-expressed in bacterial cells,while its enzymatic properties altered greatly.