大肠杆菌耐热元器件的构建及其应用

doi:10.3969/j.issn.0438-1157.2014.08.038

化工学报 ›› 2014, Vol. 65 ›› Issue (8): 3128-3135.DOI: 10.3969/j.issn.0438-1157.2014.08.038

大肠杆菌耐热元器件的构建及其应用

孙翔英¹, 刘月芹², 孙欢², 贾海洋², 戴大章², 李春^1,2

1 石河子大学化学化工学院/兵团绿色化工过程重点实验室, 新疆石河子 832003;
2 北京理工大学生命学院生物工程系, 北京 100081

收稿日期:2013-11-06 修回日期:2013-12-24 出版日期:2014-08-05 发布日期:2014-08-05
通讯作者: 李春
基金资助:
国家重点基础研究发展计划项目（2013CB733900）；国家自然科学基金项目（21376028）；博士点基金项目（20121101110050）。

Construction of heat resistance devices for Escherichia coli and their application

SUN Xiangying¹, LIU Yueqin², SUN Huan², JIA Haiyang², DAI Dazhang², LI Chun^1,2

1 Key Laboratory for Green Processing of Chemical Engineering of Xinjiang Bingtuan/School of Chemistry and Chemical Engineering, Shihezi University, Shihezi 832003, Xinjiang, China;
2 School of Life Science, Beijing Institute of Technology, Beijing 100081, China

Received:2013-11-06 Revised:2013-12-24 Online:2014-08-05 Published:2014-08-05
Supported by:
supported by the National Basic Research Program of China (2013CB733900), the National Natural Science Foundation of China (21376028) and the Doctoral Fund of Ministry of Education of China (20121101110050).

摘要/Abstract

摘要： 以提高大肠杆菌耐热性为目的，基于腾冲嗜热菌（Thermoanaerobacter tengcongensis MB4）热激蛋白基因 T.te-HSP20 构建了诱导型耐热元器件T7-T.te-HSP20 和组成型耐热元器件gapA-T.te-HSP20，转入大肠杆菌（Escherichia coli）获得工程菌 E. coli-TH 和 E. coli-GH。工程菌E. coli-TH在30℃和IPTG诱导下，目标蛋白呈可溶性表达，经50℃热激30 min后，存活率提高了3.2倍。高温发酵表明gapA-T.te-HSP20扩宽了工程菌E. coli-GH的最适生长温度的范围（37～43℃），较大程度提高了大肠杆菌的耐热性。抗逆性分析还发现工程菌E. coli-GH具备了耐热与耐丁醇的双重功能，并有一定的抗乙酸和乙醇能力。为工业梯度升温发酵生产生物基产品的高效制造、节省成本提供了新思路。

关键词: 耐热性, 热激蛋白, 存活率, 抗逆性, 大肠杆菌

Abstract: To improve the heat resistance of Escherichia coli, an inducible heat-resistance device T7-T.te-HSP20 and a constitutive heat-resistance device gapA-T.te-HSP20 based on T.te-HSP20 gene from Thermoanaerobacter tengcongensis MB4, and corresponding engineered strains E. coli-TH and E. coli-GH were constructed. The targeted protein was expressed in solubility after IPTG induction at 30℃ in E. coli-TH. Meanwhile, the survival rate of E. coli-TH was 3.2 times higher than the control at 50℃ for 30 min. The result of high-temperature fermentation showed that the optimum temperature range of E. coli-GH was broadened (37-43℃) under the regulation of heat resistance device gapA-T.te-HSP20. Stress resistance analysis showed that E. coli-GH not only possessed heat resistance and butanol resistance, but also had some resistance to acetic acid and ethanol. These results provide a new idea for modern microorganisms industry.

Key words: heat resistance, heat shock protein (HSP), survival rate, stress resistance, Escherichia coli

中图分类号:

Q789

孙翔英, 刘月芹, 孙欢, 贾海洋, 戴大章, 李春. 大肠杆菌耐热元器件的构建及其应用[J]. 化工学报, 2014, 65(8): 3128-3135.

SUN Xiangying, LIU Yueqin, SUN Huan, JIA Haiyang, DAI Dazhang, LI Chun. Construction of heat resistance devices for Escherichia coli and their application[J]. CIESC Journal, 2014, 65(8): 3128-3135.

参考文献 26

[1]	Jarne Postmus, Ronald Aardema, Leo J, de Koning. Isoenzyme expression changes in response to high temperature determine the metabolic regulation of increased glycolytic flux in yeast [J]. FEMS Yeast Research, 2012,12: 571-581
[2]	Birgit Rudolph, Katharina M Gebendorfer, Johannes Buchner, Jeannette Winter.Evolution of Escherichia coli for growth at high temperatures[J]. The Journal of Biological Chemistry, 2010, 285(25): 19029-19034
[3]	Ait-Ouazzou A, Mañas P, Condón S, Pagán R, García-Gonzalo D. Role of general stress-response alternative sigma factors σS (RpoS) and σB (SigB) in bacterial heat resistance as a function of treatment medium pH [J]. International Journal of Food Microbiology, 2012, 153: 358-364
[4]	Masayuki Murata, Hiroko Fujimoto, Kaori Nishimura. Molecular strategy for survival at a critical high temperature in E. coli [J]. PLoS One, 2011, 6(6): 1-9
[5]	Lifang Ruan, Aaron Pleitner, Michael G Gänzle, Lynn M McMullen. Solute transport proteins and the outer membrane protein NmpC contribute to heat resistance of Escherichia coli AW1.7[J]. Applied and Environmental Microbiology, 2011, 77(9): 2961-2967
[6]	Eric Guisbert, Takashi Yura, Virgil A Rhodius, Carol A Gross. Convergence of molecular, modeling, and systems approaches for an understanding of the Escherichia coli heat shock response[J]. Microbiology and Molecular. Biology Reviews, 2008, 72(3): 545-554
[7]	Chen Huayou, Chu Zhongmei, ZhangYi, Yang Shengli. Over expression and characterization of the recombinant small heat shock protein from Pyrococcus furiosus [J]. Biotechnology Letters, 2006, 28: 1089-1094
[8]	Li Dongchol, Yang Fan, Lu Bo, Chen Dianfu, Wei Junyang. Thermotolerance and molecular chaperone function of the small heat shock protein HSP20 from hyperthermophilic archaeon, Sulfolobus solfataricus P2[J]. Cell Stress and Chaperones, 2011,17: 103-108
[9]	Xue Yanfen, Xu Yi, Liu Ying, Ma Yanhe, Zhou Peijin. Thermoanaerobacter tengcongensis sp.nov., a novel anaerobic, saccharolytic, thermophilic bacterium isolated from a hot spring in Tengcong, China[J]. International Journal of Systematic and Evolutionary Microbiology, 2001, 51: 1335-1341
[10]	Hilary A Smith, Ashleigh R Burns, Tonya L Shearer, Terry W Snell. Three heat shock proteins are essential for rotifer thermotolerance[J]. Journal of Experimental Marine Biology and Ecology, 2012, 413: 1-6
[11]	James D West, Yanyu Wang, Kevin A Morano. Small molecule activators of the heat shock response: chemical properties, molecular targets, and therapeutic promise[J]. Chemical Research in Toxicology, 2012, 25: 2036-2053
[12]	Rhee Jae-Sung, Kim Ryeo-Ok, Choi Hee-Gu. Molecular and biochemical modulation of heat shock protein 20 (Hsp20) gene by temperature stress and hydrogen peroxide (H2O2) in the monogonont rotifer, Brachionus sp[J]. Comparative Biochemistry and Physiology,Part C, 2011, 154: 19-27
[13]	Kyle A, Zingaro, Eleftherios Terry Papoutsakis. GroESL over expression imparts Escherichia coli tolerance to #em/em#-, n-, and 2-butanol, 1,2,4-butanetriol and ethanol with complex and unpredictable patterns[J]. Metabolic Engineering, 2012, 7(9): 1-10
[14]	Ehrnsperger M, Gräber S, Gaestel M, Buchner J. Binding of nonnative protein to Hsp25 during heat shock creates a reservoir of folding intermediates for reactivation[J]. The EMBO Journal, 1997, 16: 221-229
[15]	Esposito L, Sica F, Sorrentino G, Berisio R, Carotenuto L, Giordano A, Raia C A, Rossi M, Lamzin V S, Wilson K S, Zagari A. Protein crystal growth in the advanced protein crystallization facility on the LMS mission: a comparison of Sulfolobus solfataricus alcohol dehydrogenase crystals grown on the ground and in microgravity[J]. Biological Crystallography, 1998, 54: 386-390
[16]	Ju Young Lee, Kyung Seok Yang, Su A Jang, Bong Hyun Sung, Sun Chang Kim. Engineering butanol-tolerance in Escherichia coli with artificial transcription factor libraries[J]. Biotechnology and Bioengineering, 2010, 108(4): 742-749
[17]	Li Yang(李阳), Tang Lixia(汤丽霞), Zheng Kai(郑楷), Wang Xiong(王雄), Jiang Rongxiang(江荣香). Optimizing expression and purification of recombinant halohydrin dehalogenase from A. radiobacter AD1[J]. CIESC Journal (化工学报), 2010, 61(12): 3213-3219
[18]	Liu Guoqiang(刘国强), Jin Hu(金虎),Gao Minjie(高敏杰),Dai Keke(戴科科),Wang Huihui(汪汇慧),Li Zhen(李震),Shi Zhongping(史仲平). Improvement of porcine interferon-A production and ATP regeneration efficiency by reducing induction temperature[J]. CIESC Journal (化工学报), 2011, 62(2): 444-450
[19]	Annette Haacke, Gabriele Fendrich, Paul Ramage. Chaperone over-expression in Escherichia coli: apparent increased yields of soluble recombinant protein kinases are due mainly to soluble aggregates[J]. Protein Expression and Purification, 2009, 64: 185-193
[20]	Jeffrey G Thomas, Francois Baneyx. Protein misfolding and inclusion body formation in recombinant Escherichia coli cells overexpressing heat-shock proteins[J]. The Journal of Biological Chemistry, 1996, 271:11141-11147
[21]	Yokoyama K, Kikuchi Y, Yasueda H. Overproduction of DnaJ in Escherichia coli improves in vivo solubility of the recombinant fish-derived transglutaminase[J]. Bioscience, Biotechnology and Biochemistry, 1998, 62(6): 1205-1210
[22]	Mary J Dunlop. Engineering microbes for tolerance to next-generation bio-fuels[J]. Biotechnol.Biofuels, 2011, 4: 32
[23]	Sergios A Nicolaou, Stefan M Gaida, Eleftherios T Papoutsakis. A comparative view of metabolite and substrate stress and tolerance in microbial bioprocessing: from biofuels and chemicals, to biocatalysis and bioremediation[J]. Metabolic. Engineering, 2010, 12(4): 307-331
[24]	Alsaker K V, Paredes C, Papoutsakis E T. Metabolite stress and tolerance in the production of biofuels and chemicals: gene-expression-based systems analysis of butanol, butyrate, and acetate stresses in the anaerobe Clostridium acetobutylicum[J]. Biotechnology and Bioengineering, 2010, 105(6): 1131-1147
[25]	Christopher A Tomas, Jeffrey Beamish, Eleftherios T Papoutsakis. Transcriptional analysis of butanol stress and tolerance in Clostridium acetobutylicum[J]. Journal of Bacteriology, 2004, 186(24): 2006-2018
[26]	James Winkler, Katy C Kao. Transcriptional analysis of Lactobacillus brevis to n-butanol and ferulic acid stress responses[J]. PLoS One, 2011, 6(8): e21438

大肠杆菌耐热元器件的构建及其应用

Construction of heat resistance devices for Escherichia coli and their application

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