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题名:
Expression and characterization of a novel metagenome-derived cellulase Exo2b and its application to improve cellulase activity in Trichoderma reesei
作者: Geng, Alei ; Zou, Gen ; Yan, Xing ; Wang, Qianfu ; Zhang, Jun ; Fanghua Liu(刘芳华) ; Zhu, Baoli ; Zhou, Zhihua
部门归属: 海岸带生物学与生物资源利用所重点实验室
通讯作者: Zhou, ZH (reprint author), Chinese Acad Sci, Key Lab Synthet Biol, Inst Plant Physiol & Ecol, Shanghai Inst Biol Sci, Fenglin Rd 300, Shanghai 200032, Peoples R China.,zhouzhihua@sippe.ac.cn
学科主题: Biotechnology & Applied Microbiology
关键词: FILAMENTOUS FUNGI ; BETA-GLUCOSIDASE ; CLOSTRIDIUM-THERMOCELLUM ; TALAROMYCES-EMERSONII ; ENZYMATIC-HYDROLYSIS ; NATURAL-PRODUCTS ; SOIL METAGENOME ; BOVINE RUMEN ; BACILLUS SP ; PURIFICATION
出处: APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
ISSN号: 0175-7598
发表日期: 2012
卷: 96, 期:4, 页:951-962
收录类别: SCI
产权排序: [Geng, Alei; Zou, Gen; Yan, Xing; Wang, Qianfu; Zhang, Jun; Liu, Fanghua; Zhou, Zhihua] Chinese Acad Sci, Key Lab Synthet Biol, Inst Plant Physiol & Ecol, Shanghai Inst Biol Sci, Shanghai 200032, Peoples R China; [Zhu, Baoli] Chinese Acad Sci, Inst Microbiol, Beijing 100101, Peoples R China
资助信息: National Basic Research Program of China (973 Program) [2011CB707403]; Chinese Academy of Sciences [KSCX1-YW-11B3, KSCX2-EW-J-12, GJHZ1128]
摘要: A metagenomic fosmid library containing 1 x 10(5) clones was constructed from a biogas digester fed with pig ordure and rice straw. In total, 121 clones with activity of 4-methylumbelliferyl-cellobiosidase were screened from the metagenomic library. A novel GH5 cellulase gene exo2b was identified from a sequenced clone EXO02C10 and expressed in Escherichia coli BL21. The corresponding recombinant Exo2b protein showed high specific activity toward both carboxymethylcellulose (CMC; 260 U/mg protein) and beta-d-glucan from barley (849 U/mg), with an optimal pH and temperature of 7.5 and 58 A degrees C, respectively. Exo2b showed stable activity at a wide pH range from 5.5 to 9.0 and was highly thermostable at 60 A degrees C in the presence of 60 mM cysteine. Residual activity was maintained at nearly 100% when Exo2b was incubated at 60 A degrees C for 15 h. A thin-layer chromatography analysis of the hydrolysis products confirmed that Exo2b was an endo-beta-1,4-glucanase and it could also produce oligosaccharide smaller than cellotetraose. The fragment encoding the Exo2b catalytic domain was then fused with the cbh1 gene from Trichoderma reesei, and the fused gene was successfully expressed in T. reesei Rut-C30. Compared to that of the parent strain, the filter paper activity and CMCase activity of the secreted proteins of a selected transformant A1 increased by 24% and 18%, respectively. Besides, the glucose concentration from the hydrolysis of pretreated corn stover by the A1 secreted proteins increased by 19.8%. The present study demonstrated the potential application of metagenome originated cellulase genes to modify cellulase producing fungi.
语种: 英语
内容类型: 共享期刊论文
URI标识: http://ir.yic.ac.cn/handle/133337/6309
Appears in Collections:科研共享资源_期刊论文

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Recommended Citation:
Geng, Alei,Zou, Gen,Yan, Xing,et al. Expression and characterization of a novel metagenome-derived cellulase Exo2b and its application to improve cellulase activity in Trichoderma reesei. 2012.
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