Abstract
D-Mannitol (hereinafter as mannitol) is a six-carbon sugar alcohol with diverse applications in food and pharmaceutical industries. To overcome the drawbacks of the chemical hydrogenation method commonly used for mannitol production at present, there is a need to search for novel prospective mannitol production strategies that are of high yield and low cost. In this study, we present a novel approach for the stoichiometric synthesis of mannitol via an in vitro synthetic enzymatic biosystem using the low-cost starch as substrate. By dividing the overall reaction pathway into three modules which could be executed sequentially in one pot, our design aimed at the stoichiometric conversion of starch-based materials into mannitol in an ATP-independent and cofactor-balanced manner. At optimized conditions, high product yields of around 95-98% were achieved using both 10 g/L and 50 g/L maltodextrin as substrate, indicating the potential of our designed system for industrial applications. This study not only provides a high-efficient strategy for the synthesis of mannitol but also expands the product scope of sugar alcohols by the in vitro synthetic enzymatic biosystems using low-cost starch-based materials as the input.
Key points
• We described a design-build-test-learn pipeline to construct in vitro biosystems.
• The designed system comprised six key enzymes and another three enzymes.
• The system converted maltodextrin stoichiometrically to mannitol in one pot.
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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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Funding
This project has received funding from the National Natural Science Foundation of China (Grant No. 21778073) and Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project (Grant No. TSBICIP-KJGG-003).
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CY conceived the synthetic enzymatic pathway. XW and CY designed the experiments, analyzed the data, and wrote the manuscript. XW, QL, and CH performed the experiments. All authors read and approved the manuscript.
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Wei, X., Li, Q., Hu, C. et al. An ATP-free in vitro synthetic enzymatic biosystem facilitating one-pot stoichiometric conversion of starch to mannitol. Appl Microbiol Biotechnol 105, 1913–1924 (2021). https://doi.org/10.1007/s00253-021-11154-9
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DOI: https://doi.org/10.1007/s00253-021-11154-9