Li Qin, Sun Baoguo, Xiong Ke, Teng Chao, Xu Youqiang, Li Liangjun, Li Xiuting
Beijing Advanced Innovation Center for Food Nutrition and Human Health, Beijing Technology and Business University (BTBU), Beijing 100048, China; School of Food and Chemical Engineering, Beijing Technology and Business University, No. 33, Fucheng Road, Beijing 100048, China.
School of Food and Chemical Engineering, Beijing Technology and Business University, No. 33, Fucheng Road, Beijing 100048, China.
Int J Biol Macromol. 2017 Mar;96:451-458. doi: 10.1016/j.ijbiomac.2016.12.050. Epub 2016 Dec 21.
The purpose of this study was to gain insights into the hydrolysis characteristics of xylanase in producing xylo-oligosaccharides and to improve these by protein engineering. In this study, a hybrid enzyme (T-XynFM) was created from Talaromyces thermophilus F1208 xylanase (T-Xyn) by replacement of the N-terminal extension Phe1-Pro16 with Ala1-Gln8 and substitution of amino acid Phe193 (185) with Ser in the C-terminal. The enzyme showed optimum activity at 55°C and pH 6.5. Its residual activity was more than 55% after treatment at 50°C, pH 6.5 for 12h Km values of T-XynFM for beechwood xylan, birchwood xylan, and oat-spelt xylan were 10.31, 10.03, and 8.90mgmL, respectively. The enzyme displayed special hydrolysis characteristics: almost no xylose was produced on using xylotriose (X3), xylotetraose (X4), and xylopentaose as substrates. Moreover, over 60% X4 existed in hydrolyzed products of X3, indicating that T-XynFM possesses formidable transglycosylation properties.
本研究的目的是深入了解木聚糖酶在生产低聚木糖中的水解特性，并通过蛋白质工程对其进行改进。在本研究中，通过将嗜热栖热菌F1208木聚糖酶（T-Xyn）的N端延伸序列Phe1-Pro16替换为Ala1-Gln8，并将C端的氨基酸Phe193（185）替换为Ser，构建了一种杂合酶（T-XynFM）。该酶在55°C和pH 6.5时表现出最佳活性。在50°C、pH 6.5条件下处理12小时后，其残余活性超过55%。T-XynFM对山毛榉木聚糖、桦木木聚糖和燕麦-斯佩尔特木聚糖的Km值分别为10.31、10.03和8.90mg/mL。该酶表现出特殊的水解特性：以木三糖（X3）、木四糖（X4）和木五糖为底物时几乎不产生木糖。此外，X3水解产物中X4的含量超过60%，表明T-XynFM具有强大的转糖基化特性。
