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Functional Characterization of Recombinant Endo-Levanase (LevBk) from Bacillus koreensis HL12 on Short-Chain Levan-Type Fructooligosaccharides Production

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Abstract

Levan-type fructooligosaccharides (L-FOSs) are a prominent class of non-digestible oligosaccharides with potential as nutritional prebiotics. Endo-levanase, which randomly hydrolyzes β-(2,6)-linkages in fructans, is a promising enzyme for short-chain FOS production. In this work, a recombinant levanase (LevBk) from Bacillus koreensis strain HL12 was characterized. Soluble LevBk protein was produced in Escherichia coli BL21(DE3) system at 40 mg/L of culture medium. Based on sequence and structural analysis, LevBk was classified as a member of endo-levanase in GH32 family containing N-terminal substrate binding pocket and C-terminal β-sandwich domains. LevBk optimally worked at 45 °C, pH 6.0 with the specific activity of 2.43 U/mg. Based on enzymatic hydrolysis, short-chain L-FOSs with degree of polymerization (DP) of 2–4 were produced from hydrolysis of timothy grass levan under optimal conditions for 9–24 h. With its ability to produce L-FOSs with specific chain lengths, LevBk could be attractively applied for converting of levan containing material to high value-added sweetener in the biorefinery industry.

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Data Availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

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Acknowledgements

The authors gratefully acknowledge the financial support by Faculty of Science and Technology, Thammasat University, Contract No SciGR 9/2564. The authors acknowledge the support provided by Enzyme Technology Research Team, National Center for Genetic Engineering and Biotechnology and Department of Biotechnology, Faculty of Science and Technology, Thammasat University. The authors would like to thank Dr. Nattapol Arunrattanamook for manuscript proofreading and comments.

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Authors and Affiliations

  1. Department of Biotechnology, Faculty of Science and Technology, Rangsit Campus, Thammasat University, Pathum Thani, 12120, Thailand

    Hataikarn Lekakarn, Phuphiphat Jaikaew & Thanyanun Kuantum

  2. Enzyme Technology Research Team, Biorefinery Technology and Bioproduct Research Group, National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Phahonyothin Road, Khlong Luang, Pathum Thani, 12120, Thailand

    Benjarat Bunterngsook & Verawat Champreda

  3. Cassava and Starch Technology Research Team, Functional Ingredients and Food Innovation Research Group, National Center for Genetic Engineering and Biotechnology, Bangkok, 10900, Thailand

    Rungtiva Wansuksri

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Contributions

HL, BB and VC conceived and designed the work. HL, BB, TK, RW and PJ performed the experiments, and analyzed the data. HL, BB, PJ and RW drafted the manuscript. BB and VC critically revised the manuscript for intellectual content. All authors have read and agree to the submission of the manuscript.

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Correspondence to Benjarat Bunterngsook.

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Lekakarn, H., Bunterngsook, B., Jaikaew, P. et al. Functional Characterization of Recombinant Endo-Levanase (LevBk) from Bacillus koreensis HL12 on Short-Chain Levan-Type Fructooligosaccharides Production. Protein J 41, 477–488 (2022). https://doi.org/10.1007/s10930-022-10069-2

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