CAJ | 학술논문

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。
장래자대장간균K-12적L-아랍백당이구매기인araA정향삽입표체재체pET-32a （+）병전입대장간균BL21（DE3）중。경PCR、쌍매절감정화서렬분석，증실중조질립pET32a-araA구건성공。중조균경isopropylthio-β-D-galactoside（IPTG）유도표체목적단백，주요이가용형식존재。이중현균액위매원，D-반유당위저물，재온도60℃、pH7.6시매활최대，가입1 mmol/L Mn2+가사활력제고50%이상。
@@@@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 Mn2+ ions enhanced the enzyme activity. In the presence of 1 mmol/L Mn2+ions,the relative activity increased by 50%.