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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

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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 N-terminal-fused signal peptides. We found that the matured XynHB itself could be auto-secreted 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.

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Acknowledgments

LY and CY acknowledge the support from State Key Laboratory of Biocatalysis and Enzyme Engineering.

Funding

This work was supported by the National Key Technology R & D Program of China (2018YFD0500203 to LY, CY, and LF) and the National Natural Science Foundation of China (No. 31870057 to LY).

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Authors and Affiliations

  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

    Faying Zhang, Huahua He, Ting Deng, Haoran Ge, Chan Yu & Li Yi

  2. School of Environmental Studies, China University of Geosciences, Wuhan, 430074, China

    Liang Feng

  3. Hubei Key Laboratory of Lipid Chemistry and Nutrition, Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Wuhan, 430062, China

    Fenghong Huang

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  1. Faying Zhang
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  4. Haoran Ge
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  8. Li Yi
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Contributions

FZ, HH, TD, and HG conducted the experiments. FH, LF, and CY contributed the conceptualization. FZ, HH, and LY analyzed the results. LY conceived the idea for the project and wrote the paper with FZ and HH.

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Correspondence to Li Yi.

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Zhang, F., He, H., Deng, T. et al. N-Terminal Fused Signal Peptide Prompted Extracellular Production of a Bacillus-Derived Alkaline and Thermo Stable Xylanase in E. coli Through Cell Autolysis. Appl Biochem Biotechnol 192, 339–352 (2020). https://doi.org/10.1007/s12010-020-03323-9

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