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Biosensor-informed engineering of Cupriavidus necator H16 for autotrophic D-mannitol production

Hanko, Erik K.R.; Sherlock, Gillian; Minton, Nigel P.; Malys, Naglis

Authors

Erik K.R. Hanko

Gillian Sherlock

Naglis Malys



Abstract

Cupriavidus necator H16 is one of the most researched carbon dioxide (CO2)-fixing bacteria. It can store carbon in form of the polymer polyhydroxybutyrate and generate energy by aerobic hydrogen oxidation under lithoautotrophic conditions, making C. necator an ideal chassis for the biological production of value-added compounds from waste gases. Despite its immense potential, however, the experimental evidence of C. necator utilisation for autotrophic biosynthesis of chemicals is limited. Here, we genetically engineered C. necator for the high-level de novo biosynthesis of the industrially relevant sugar alcohol mannitol directly from Calvin-Benson-Bassham (CBB) cycle intermediates. To identify optimal mannitol production conditions in C. necator, a mannitol-responsive biosensor was applied for screening of mono- and bifunctional mannitol 1-phosphate dehydrogenases (MtlDs) and mannitol 1-phosphate phosphatases (M1Ps). We found that MtlD/M1P from brown alga Ectocarpus siliculosus performed overall the best under heterotrophic growth conditions and was selected to be chromosomally integrated. Consequently, autotrophic fermentation of recombinant C. necator yielded up to 3.9?g/L mannitol, representing a substantial improvement over mannitol biosynthesis using recombinant cyanobacteria. Importantly, we demonstrate that at the onset of stationary growth phase nearly 100% of carbon can be directed from the CBB cycle into mannitol through the glyceraldehyde 3-phosphate and fructose 6-phosphate intermediates. This study highlights for the first time the potential of C. necator to generate sugar alcohols from CO2 utilising precursors derived from the CBB cycle.

Citation

Hanko, E. K., Sherlock, G., Minton, N. P., & Malys, N. (2022). Biosensor-informed engineering of Cupriavidus necator H16 for autotrophic D-mannitol production. Metabolic Engineering, 72, 24-34. https://doi.org/10.1016/j.ymben.2022.02.003

Journal Article Type Article
Acceptance Date Feb 7, 2022
Online Publication Date Feb 8, 2022
Publication Date Jul 1, 2022
Deposit Date Feb 14, 2022
Publicly Available Date Feb 9, 2023
Journal Metabolic Engineering
Print ISSN 1096-7176
Electronic ISSN 1096-7184
Publisher Elsevier
Peer Reviewed Peer Reviewed
Volume 72
Pages 24-34
DOI https://doi.org/10.1016/j.ymben.2022.02.003
Keywords Applied Microbiology and Biotechnology; Bioengineering; Biotechnology
Public URL https://nottingham-repository.worktribe.com/output/7466577
Publisher URL https://www.sciencedirect.com/science/article/pii/S1096717622000271

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