黑曲霉内切β-1,3(4)-葡聚糖酶的基因克隆与酶学特性分析  

Gene cloning and biochemical characterization of endo-1,3(4)-β-glucanases from Aspergillus niger

作　　者：董自星[1] 李伟国[2] 佟新新 田康明[1] 路福平[2] 

机构地区：[1]天津科技大学化工与材料学院,天津300457 [2]天津科技大学生物工程学院,天津300457

出　　处：《食品与发酵工业》2016年第11期58-64,共7页Food and Fermentation Industries

摘　　要：基于黑曲霉CBS 513.88的基因组信息,以黑曲霉CICIM F0510为出发菌株,提取其总RNA,反转录得到c DNA,成功将其3个内切β-1,3(4)-葡聚糖酶的基因(en3g A、en3g B、en3g C)在毕赤酵母中进行了克隆与表达。获得了3个重组菌GS115(p PIC-en3g A)、GS115(p PIC-en3g B)和GS115(p PIC-en3g C)。在摇瓶水平上,这株重组菌的酶活分别为1.3、8.5和10.1 U/m L。重组酶En3g A、En3g B和En3g C的最适作用温度分别为65、50和55°C;最适作用p H分别为5.0、5.0和3.5。此外,3种重组酶对羧甲基纤维素钠(sodium carboxymethylcellulose,CMC-Na)和凝结多糖都具有典型的内切水解作用。这3个重组β-1,3(4)-葡聚糖酶具有良好的温度和p H稳定性,可为其在啤酒制造以及饲料加工过程中的应用奠定基础。Based on the genome information of Aspergillus niger CBS 513.88, total RNA was extracted from A. niger CICIM F0510 and reverse transcribed to cDNA. Using the cDNA as a template, three genes encoding endo-1,3 （4）-β-glucanases （en3gA, en3gB and en3gC） were successfully cloned and expressed in Pichia pastoris GSll5. Three recombinant stained named as GS115 （pPIC-en3gA） , GS115 （pPIC-en3gB） and GS115 （pPIC-en3gC） were obtained. At the shake-flask fermentation level, the activities of these recombinant endo-1,3 （4） -β-glucanases were determined to be 1.3, 8.5 and 10.1 U/mL, respectively. The optimal reaction temperatures for recombinant enzymes En3gA, En3gB and En3gC were 65, 50 and 55 ℃ , respectively. Their optimal reaction pH values were 5.0, 5.0 and 3.5, respectively. In addition, these three recombinant enzymes also hydrolyzed internal bonds in sodium car- boxymethylcellulose （CMC-Na） and curdlan. Their distinct biochemical properties will lay a solid foundation for their applications in brewing and feed manufacture.

关 键 词：内切β-1 3(4)-葡聚糖酶 黑曲霉 酶学特征 

分 类 号：TQ925[轻工技术与工程—发酵工程]