木聚糖酶是一种新型的安全无毒面粉改良剂,能有效改善面团性能和烘焙质量。为了挖掘新的木聚糖酶资源,并评估其在面包烘焙中的应用,该研究在大肠杆菌中融合Sumo标签重组表达了来自嗜热微生物Caldicellulosiruptor owensensis的木聚糖酶,并对融合木聚糖酶SumoXyn的酶学性质和对面包烘焙的品质影响进行了初步研究。大肠杆菌能高效表达C.owensensis的木聚糖酶,且仅需通过75 ℃加热细胞破碎液30 min这种简单操作即可得到纯度较高的融合木聚糖酶SumoXyn,微量差示扫描荧光分析发现该酶热变性中点温度值为83.5 ℃。对其酶学性质研究发现该酶最适pH值为9.0,但在pH 5~10时仍有60%以上的活性。在pH 4~10的缓冲液中处理24 h,SumoXyn仍有80%以上的残余活性;该酶在80 ℃时酶活性最高,但在100 ℃仍有50%的酶活性,75 ℃加热1 h后仍有约75%的残余活性,表明SumoXyn具有较好的pH稳定性和热稳定性。以榉木木聚糖为底物,测得其vmax为2 354.52 μmol/(min·mg),Km为2.26 mg/mL,Kcat为214.68 s-1。以200 g高筋全麦面粉为原料制作面包,研究添加量分别为3、6、9 mg的SumoXyn对全麦面包品质的影响,发现随着SumoXyn添加量的增加,面包比容和弹性增加、硬度和咀嚼性降低,全麦面包品质得到显著改善,说明SumoXyn是一种具有潜力的面粉改良剂。
Xylanase is a novel and safe flour additive that effectively enhances dough properties and baking quality.To explore new sources of xylanase and evaluate its application in bread baking, xylanase from Caldicellulosiruptor owensensis was heterogeneously expressed in Escherichia coli by fusion with Sumo tag.The enzymatic properties of the xylanase SumoXyn and its impact on bread quality were investigated.The xylanase from Caldicellulosiruptor owensensis was efficiently expressed in E.coli and the enzyme was obtained with high purity by simply heating the cell lysate at 75 ℃ for 30 minutes.Nano DSF analysis showed that the Tm value of this enzyme was 83.5 ℃.The optimum pH of the enzyme was 9.0, but it still had more than 60% activity at pH 5-10.Treated in pH 4-10 buffer for 24 h, SumoXyn still had more than 80% residual activity.SumoXyn displayed the highest activity at 80 ℃, but it still has 50% of the highest activity at 100 ℃, and the residual activity was about 75% after heating at 75 ℃ for 1 h, indicating that Sumoxyn had good pH stability and thermal stability.Using bagasse xylan as substrate, the Vmax was 2 354.52 μmol/(min·mg), the Kmwas 2.26 mg/mL and the Kcat was 214.68 S-1.Using 200 g high gluten whole wheat flour to make bread, the effects of SumoXyn supplemented with 3 mg, 6 mg, 9 mg, respectively, on the quality of whole wheat bread were investigated.It was found that with the increase of SumoXyn supplemented, the specific volume and elasticity of bread increased, the hardness and chewability decreased, and the quality of whole wheat bread was significantly improved.These findings suggest that SumoXyn has great potential as a flour improver.
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