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d-Allulose Production from d-fructose by Putative Dolichol Phosphate Mannose Synthase from Bacillus sp. with Potential d-allulose 3-epimrase Activity

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Abstract

We found a putative dolichol phosphate mannose synthases (DPMS) from Bacillus sp., which exhibited the highest specific activity toward d-allulose, one of a functional rare sugars and also known as d-psicose, and identified it as d-allulose 3-epimerase (Basp_DAEase). The gene of Basp_DAEase from Bacillus sp. was cloned to the expression vector (pET-28a(+)) and then expressed as a recombinant enzyme in the Escherichia coli BL21(DE3). The recombinant enzyme was purified homogeneously on the SDS-PAGE with 34 kDa by 6XHis-tagging affinity chromatography and it was found to exist in dimer form as its activity form by Sephacryl S 200 HR 16/60 gelfiltration chromatography. The reaction conditions for a recombinant Basp_DAEase were optimized to produce d-allulose from d-fructose. The epimerization activity of Basp_DAEase toward d-fructose was maximum at pH 7.5 and 40°C with 1 mM Co2+. The half-lives of Basp_DAEase at 35°C, 40°C, 45°C, 50°C, and 55°C were 45.8, 4.18, 1.24, 0.46, and 0.17 h, respectively. At pH 7.5, 40°C, and 1mM Co2+, 215 g/L of d-allulose was produced from 700 g/L of d-fructose by 20 U/mL of Basp_DAEase after 50 min of enzyme reaction with a conversion yield of 31% and productivity of 258 g/L/h. This is the highest productivity of d-allulose from d-fructose reported to date. From this result, it is strongly suggested that Basp_DAEase has high potential application value in the industrial d-allulose production from d-fructose.

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Acknowledgements

This study was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (MSIT) (No. 2019R1F1A1059906) and R&D Program for Forest Science Technology (Project No. 2020197A00-2022-BA01) provided by the Korea Forest Service (Korea Forestry Promotion Institute).

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  1. Min-Ju Seo and Eun Ryeong Kwon have contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, St. Paul, Minneapolis, MN, 55108, USA

    Min-Ju Seo

  2. Department of Food Science and Technology, Catholic University of Daegu, Gyeongsan, 38430, Korea

    Eun Ryeong Kwon, Sang Jin Kim, Min Su Choi & Chang-Su Park

  3. Division of Wild Plat Industrialization Research, Baekdudaegan National Arboretum, Bonghwa, 26209, Korea

    Yeong-Su Kim

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  1. Min-Ju Seo
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Correspondence to Chang-Su Park.

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Seo, MJ., Kwon, E.R., Kim, S.J. et al. d-Allulose Production from d-fructose by Putative Dolichol Phosphate Mannose Synthase from Bacillus sp. with Potential d-allulose 3-epimrase Activity. Biotechnol Bioproc E 26, 976–984 (2021). https://doi.org/10.1007/s12257-021-0007-3

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