caa a22 4500 605505683 CHVBK 20210128100623.0 cr unu---uuuuu 210128e20151101xx s 000 0 eng 10.1007/s00253-015-6619-9 doi (NATIONALLICENCE)springer-10.1007/s00253-015-6619-9 Overexpression and characterization of a glucose-tolerant β-glucosidase from T. aotearoense with high specific activity for cellobiose [Elektronische Daten] [Fang Yang, Xiaofeng Yang, Zhe Li, Chenyu Du, Jufang Wang, Shuang Li] Thermoanaerobacterium aotearoense P8G3#4 produced β-glucosidase (BGL) intracellularly when grown in liquid culture on cellobiose. The gene bgl, encoding β-glucosidase, was cloned and sequenced. Analysis revealed that the bgl contained an open reading frame of 1314bp encoding a protein of 446 amino acid residues, and the product belonged to the glycoside hydrolase family 1 with the canonical glycoside hydrolase family 1 (GH1) (β/α)8 TIM barrel fold. Expression of pET-bgl together with a chaperone gene cloned in vector pGro7 in Escherichia coli dramatically enhanced the crude enzyme activity to a specific activity of 256.3U/mg wet cells, which resulted in a 9.2-fold increase of that obtained from the expression without any chaperones. The purified BGL exhibited relatively high thermostability and pH stability with its highest activity at 60°C and pH 6.0. In addition, the activities of BGL were remarkably stimulated by the addition of 5mM Na+ or K+. The enzyme showed strong ability to hydrolyze cellobiose with a K m and V max of 25.45mM and 740.5U/mg, respectively. The BGL was activated by glucose at concentration varying from 50 to 250mM and tolerant to glucose inhibition with a K i of 800mM glucose. The supplement of the purified BGL to the sugarcane bagasse hydrolysis mixture containing a commercial cellulase resulted in about 20% enhancement of the released reducing sugars. These properties of the purified BGL should have important practical implication in its potential applications for better industrial production of glucose or bioethanol started from lignocellulosic biomass. Springer-Verlag Berlin Heidelberg, 2015 β-Glucosidase nationallicence Thermoanaerobacterium aotearoense P8G3#4 nationallicence Glucose tolerance nationallicence Cellobiose degradation nationallicence Chaperones nationallicence Yang Fang Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut Yang Xiaofeng Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut Li Zhe Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut Du Chenyu School of Applied Sciences, The University of Huddersfield, Queensgate, HD1 3DH, Huddersfield, UK aut Wang Jufang Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut Li Shuang Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/21(2015-11-01), 8903-8915 0175-7598 99:21<8903 2015 99 253 https://doi.org/10.1007/s00253-015-6619-9 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6619-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Yang Fang Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Yang Xiaofeng Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Li Zhe Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Du Chenyu School of Applied Sciences, The University of Huddersfield, Queensgate, HD1 3DH, Huddersfield, UK aut NATIONALLICENCE 700 1- Wang Jufang Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut NATIONALLICENCE 700 1- Li Shuang Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Bioscience and Bioengineering, South China University of Technology, 510006, Guangzhou, China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/21(2015-11-01), 8903-8915 0175-7598 99:21<8903 2015 99 253