摘要
将来自大肠杆菌K-12的L-阿拉伯糖异构酶基因araA定向插入表达载体pET-32a(+)并转入大肠杆菌BL21(DE3)中。经PCR、双酶切鉴定和序列分析,证实重组质粒pET32a-araA构建成功。重组菌经isopropylthio-β-D-galactoside(IPTG)诱导表达目的蛋白,主要以可溶形式存在。以重悬菌液为酶源,D-半乳糖为底物,在温度60℃、pH7.6时酶活最大,加入1 mmol/L Mn2+可使活力提高50%以上。
The gene araA encoding L-arabinose isomerase from Escherichia coli K12 was cloned into expression vector pET-32a. Then the recombinant plasmid was transformed into strain E. coli BL21 （DE3）. Target proteins L-arabinose isomerase was highly expressed with IPTG. Analysis by SDS-PAGE showed that the target enzymes were expressed in soluble form. To investigate the properties of L-arabinose isomerase, D-galactose was used as substrate and resuspended cells as enzyme source.The results showed that the optimal temperature and pH of the enzyme were 60 ℃ and 7.6,respectively. The addition of Mn^2＋ ions enhanced the enzyme activity. In the presence of 1 mmol/L Mn^2＋ ions,the relative activity increased by 50 %.
出处
《食品研究与开发》
CAS
北大核心
2013年第11期68-71,共4页
Food Research and Development
