Thermus thermophilus分支酶的重组表达及其在抗性糊精制备中的应用
Recombinant expression and application in preparation of resistant dextrin of branching enzyme from Thermus thermophilus
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摘要: 本文研究了来源于Thermus thermophilus的淀粉分支酶TtSBE,通过基因工程的方法构建重组大肠杆菌E.coli BL21（PET-24（a+）-TtSBE）,使之在大肠杆菌内高效表达,并在此基础上对TtSBE的酶学性质进行研究,结果表明,TtSBE的最适pH为6.5,最适温度为60℃,在60℃的半衰期为40 h左右,TtSBE经纯化后,比活为773.27 U/mg;以2%的焦糊精为底物,利用TtSBE制备抗性糊精,当酶转化温度为60℃,pH为6.5,加酶量为3000 U/g（焦糊精）,酶转化时间为12 h时,抗性成分含量达到50%,较焦糊精原料中的抗性成分含量提高了7%,本研究为抗性糊精的产率和质量的进一步提高奠定了基础。Abstract: In this paper, through the construction of recombinant E.coli BL21 ( PET-24 ( a +) -TtSBE) by genetic engineering method, the starch branching enzyme TtSBE from Thermus thermophilus was overexpressed.The enzymatic properties of TtSBE were studied.The results showed that the optimum pH of TtSBE was 6.5, the optimum temperature was 60 ℃, the half-life at 60 ℃ was about 40 h, and the specific activity of TtSBE was 773.27 U/mg after purification. By using 2% pyrodextrin as substrate, the TtSBE conversion temperature 60 ℃, pH6.5, the amount of enzyme 3000 U/g ( pyrodextrin) , after treated for 12 h, the content of the resistant component reached 50%, which was 7% higher than raw material. This study would lay the foundation for the further improvement of yield and quality of resistant dextrin.
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