N-Terminal Fused Signal Peptide Prompted Extracellular Production of a Bacillus- Derived Alkaline and Thermo Stable Xyla
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N-Terminal Fused Signal Peptide Prompted Extracellular Production of a Bacillus-Derived Alkaline and Thermo Stable Xylanase in E. coli Through Cell Autolysis Faying Zhang 1 & Huahua He 1 & Ting Deng 1 & Haoran Ge 1 & Chan Yu 1 & Liang Feng 2 & Fenghong Huang 3 & Li Yi 1 Received: 6 January 2020 / Accepted: 23 April 2020/ # Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract
Xylanases are extensively used as industrial enzymes for its ability of hydrolyzing xylan to oligosaccharides. Here, XynHB, a thermo and alkaline stable xylanase derived from Bacillus pumilus HBP8, was extracellularly produced in E. coli cells through Nterminal-fused signal peptides. We found that the matured XynHB itself could be autosecreted out of E. coli BL21(DE3) cells at a very low level, and two Sec-pathway signal peptides, PelB and OmpA, and one dual Sec-Tat-pathway signal peptide, FhuD, could effectively prompt its extracellular production up to 12-fold. Our results showed that PelB signal peptide led to the highest extracellular production of XynHB for approximately 54.1 μg/mL, and FhuD-fused XynHB possessed the highest specific activity of 1746.0 U/ mg at 70 °C. Meanwhile, our studies also indicated that PelB- and FhuD-fused XynHB might disrupt E. coli cells’ periplasm during their secretion process, thus causing cell lysis to facilitate their extracellular production. Moreover, further characterization revealed that the extracellular production of XynHB was not affected by the outer membrane permeability of E. coli cells. Our studies provided an advantageous strategy for the extracellular production of xylanase in E. coli, which may also be used for E. coli autolysis in the future. Keywords Xylanase . Extracellular production . Secretion . Signal peptides . Cell autolysis
* Li Yi [email protected]
1
State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Engineering Research Center for Bio-enzyme Catalysis, College of Life Sciences, Hubei University, NO. 368 Youyi Road, Wuchang District, Wuhan 430062 Hubei, China
2
School of Environmental Studies, China University of Geosciences, Wuhan 430074, China
3
Hubei Key Laboratory of Lipid Chemistry and Nutrition, Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Wuhan 430062, China
Applied Biochemistry and Biotechnology
Introduction Endo-β-1,4-xylanases (EC.3.2.1.8) exclusively catalyze the hydrolysis of internal β-1,4-Dxylosidic linkages of xylan to produce oligosaccharides via a double displacement mechanism, which could be subsequently hydrolyzed to xylose by β-xylosidase [1]. As the major component of hemicellulose and the second most abundant renewable polysaccharide in nature, bio-catalysis of xylan by xylanase could bring great profits in industries, including textile, kraft bleaching, animal feed, bakery, and green energy production [2–4]. Considering its wide range of industrial applications, xylanases with fine properties and their effective large-scale production are constantly of great interests. Heretofore
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