松二糖具有低热量和不致龋齿的特性,在食品工业具有广泛的应用前景。与淀粉蔗糖酶相比,Caulobacter crescentus蔗糖水解酶突变体S271A (CcSH S271A)转化蔗糖过程中产生的麦芽寡糖副产物少,可代替淀粉蔗糖酶用于松二糖制备。该研究将CcSH S271A在枯草芽孢杆菌(Bacillus subtilis)中重组表达,对该重组酶进行酶学表征并对其制备松二糖的工艺进行了优化。结果表明,重组酶的酶促反应最适温度为45 ℃,最适pH为8.0,在30 ℃时热稳定性好,半衰期为312 h。当底物蔗糖浓度为2.0 mol/L,温度30 ℃、pH 8.0、加酶量为60 U/g蔗糖,反应时间48 h时,松二糖最高产率达到66.3%。是以蔗糖为单一底物制备松二糖的最高水平,为松二糖的制备奠定了基础。
Turanose has the characteristics of low calorie and non-caries properties, so it has a wide application prospect in food industry. Compared with amylosucrase, the Caulobacter crescentus sucrose hydrolase mutant S271A (CcSH S271A) produced fewer malto-oligosaccharide by-products, which can replace amylosucrase for the preparation of turanose. In this study, the CcSH S271A was recombinantly expressed in Bacillus subtilis, the enzymatic properties of recombinant CcSH S271A were identified and the process of preparing turanose by recombinant CcSH S271A was optimized. The results showed that the optimal reaction temperature and reaction pH of recombinant CcSH S271A were 45 ℃ and 8.0, respectively. The recombinant CcSH S271A displayed excellent thermal stability at 30 ℃ with a half-life of 312 h. The yield of turanose was 66.3% with 2 mol/L sucrose as the substrate, when the amount of enzyme was 60 U/g sucrose, at pH 8.0 and 30 ℃ for 48 h. It is the highest level of preparing turanose using sucrose as a single substrate, which lays a solid foundation for the preparation of turanose.
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