Cloning, expression and characterization of a cold-adapted endo-1, 4-beta-glucanase from Citrobacter farmeri A1, a symbiotic bacterium of Reticulitermes labralis

文献类型： 外文期刊

第一作者： Bai, Xi

作者： Bai, Xi;Yuan, Xianjun;Li, Junfeng;Shao, Tao;Wen, Aiyou;Bai, Yunfeng

作者机构：

关键词： Citrobacter farmeri;Endoglucanase;Cold-adapted;Expression;Escherichia coli;Properties

期刊名称：PEERJ （ 影响因子：2.984； 五年影响因子：3.369 ）

ISSN： 2167-8359

年卷期： 2016 年 4 卷

页码：

收录情况： SCI

摘要： Background. Many biotechnological and industrial applications can benefit from cold-adapted EglCs through increased efficiency of catalytic processes at low temperature. In our previous study, Citrobacter farmeri A1 which was isolated from a wood-inhabiting termite Reticulitermes labralis could secrete a cold-adapted EglC. However, its EglC was difficult to purify for enzymatic properties detection because of its low activity (0.8 U/ml). The objective of the present study was to clone and express the C. farmeri EglC gene in Escherichia coli to improve production level and determine the enzymatic properties of the recombinant enzyme. Methods. The EglC gene was cloned from C. farmeri A1 by thermal asymmetric interlaced PCR. EglC was transformed into vector pET22b and functionally expressed in E. coli. The recombination protein EglC22b was purified for properties detection. Results. SDS-PAGE revealed that the molecular mass of the recombinant endoglucanase was approximately 42 kDa. The activity of the E. coli pET22b-EglC crude extract was 9.5 U/ml. Additionally, it was active at pH 6.58.0 with an optimum pH of 7.0. The recombinant enzyme had an optimal temperature of 3040 degrees C and exhibited >50% relative activity even at 5 degrees C, whereas it lost approximately 90% of its activity after incubation at 60 degrees C for 30 min. Its activity was enhanced by Co2+ and Fe3+, but inhibited by Cd2+, Zn2+, Li+, Triton X-100, DMSO, acetonitrile, Tween 80, SDS, and EDTA. Conclusion. These biochemical properties indicate that the recombinant enzyme is a cold-adapted endoglucanase that can be used for various industrial applications.

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