东方肉座菌EU7-22木聚糖酶和木糖苷酶基因克隆及生物信息学研究  

Gene cloning and bioinformatics analysis of Xylanases and Xylosidase from Hypocrea orientalis

作　　者：龙传南[1] 蒋凤姣[1] 邬小兵[2] 刘健[1] 龙敏南[1,2] 

机构地区：[1]厦门大学能源研究院,福建厦门361005 [2]厦门大学生命科学学院,福建厦门361005

出　　处：《生物信息学》2013年第1期58-64,共7页Chinese Journal of Bioinformatics

基　　金：国家重点基础研究发展计划(973计划)(NO.2010CB732201);中央高校基本科研业务费专项资金(NO.201112G026);国家自然科学基金(NO.31170067)

摘　　要：东方肉座菌EU7-22具有高产半纤维素酶的能力。根据已报道的同属里氏木霉及绿色木霉木聚糖酶,木糖苷酶相关基因序列,设计PCR引物扩增出东方肉座菌内切木聚糖酶(XYNⅠ,XYNⅡ)及β-木糖苷酶(β-BXL)基因。序列经NCBIBlast分析:东方肉座菌xynⅠ基因与里氏木霉xyn1基因(X69573.1)的同源性最高达到91%;xynⅡ基因与绿色木霉xyn2基因(EF079061)同源性最高达到93%;β-bxl基因与里氏木霉β-bxl1基因(Z69257.1)的同源性最高达到94%。生物信息学分析表明内切木聚糖酶Ⅰ和Ⅱ均属于糖基水解酶家族11,N末端前19个氨基酸均为信号肽。内切木聚糖酶Ⅰ分子量为24.13kD,等电点为7.87,含有3个糖基化位点;内切木聚糖酶Ⅱ分子量为24.44kD,等电点为4.86,含有1个糖基化位点;β-木糖苷酶属于糖基水解酶家族3,分子量为87.27kD,等电点为5.49,N末端前20个氨基酸为信号肽,含有8个糖基化位点。利用SWISS-Model对木聚糖酶,木糖苷酶蛋白质三级结构进行了预测和模拟。对木聚糖酶和木糖苷酶基因及其编码蛋白质的生物信息学分析,为进一步研究这些基因的表达与调控、构建高效利用纤维素组份的工程菌株奠定基础。Hypocrea orientalis EU7 - 22 had a high potential to yield hemicellulase. Two endo - xylanases genes （ xyn I, xyn I1 ） and one β- xylosidase gene （ β- bxl） were successfully cloned by PCR, according to the repor- ted xylanases and β-xylosidase gene sequences of Trichoderma reesei and Trichoderma viride. The xyn I and β- bxl gene from H. orientalis showed the highest nucleotide homology of 91% and 94% with the corresponding gene from T. reesei, respectively. While the xyn II gene from H. orientalis showed the highest of 93 % nucleotide homol- ogy with the corresponding gene from T. viride. The bioinformatics analysis indicated that the enzyme XYN I and XYN II belonged to the glycosyl hydrolase family 11 with the molecular weight of 24.13 kD and 24.44 kD, respec- tively. The first 19 AA of N - terminal of XYN I and XYN II were the signal peptide sequence. The enzyme XYN I and XYN II contained three and one N - glycosylation site, respectively. The isoelectric point of enzyme XYN I and XYN II were identified as 7.87 and 4.86, respectively. The enzyme β- BXL belonged to glycosyl hydrolase family 3 with molecular weight of 87.27 kD and isoelectric point of 5.49. The first 20 AA of N - terminal of β - BXL belonged to signal peptide sequence. The enzyme contained 8 N - glycosylation sites. By using SWISS - Model, the tertiary structure of the three enzyme proteins were predicted and simulated. These genes cloning and bioin- formatics analysis would help to the further research on mechanism of expression and regulation of hemicellulase.

关 键 词：东方肉座菌 木聚糖酶 木糖苷酶 基因克隆 生物信息学 

分 类 号：Q343.15[生物学—遗传学]