Abstract
Gluconobacter is a potential strain for single-step production of 2-keto-L-gulonic acid (2-KLG), which is the direct precursor of vitamin C. Three dehydrogenases, namely, sorbitol dehydrogenase (SLDH), sorbose dehydrogenase (SDH), and sorbosone dehydrogenase (SNDH), are involved in the production of 2-KLG from D-sorbitol. In the present study, the potential SNDH/SDH gene cluster in the strain Gluconobacter cerinus CGMCC 1.110 was mined by genome analysis, and its function in transforming L-sorbose to 2-KLG was verified. Proteomic analysis showed that the expression level of SNDH/SDH had a great influence on the titer of 2-KLG, and fermentation results showed that SDH was the rate-limiting enzyme. A systematic metabolic engineering process, which was theoretically suitable for increasing the titer of many products involving membrane-bound dehydrogenase from Gluconobacter, was then performed to improve the 2-KLG titer in G. cerinus CGMCC 1.110 from undetectable to 51.9 g/L in a 5-L bioreactor after fermentation optimization. The strategies used in this study may provide a reference for mining other potential applications of Gluconobacter.
Key points
• The potential SNDH/SDH gene cluster in G. cerinus CGMCC 1.110 was mined.
• A systematic engineering process was performed to improve the titer of 2-KLG.
• The 2-KLG titer was successfully increased from undetectable to 51.9 g/L.
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Data availability
The data that support the findings of this study are available from the corresponding author upon reasonable request.
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Funding
This work was supported by the National Natural Science Foundation of China (Key Program, 31830068), the Foundation for Innovative Research Groups of the National Natural Science Foundation of China (32021005), and the National Science Fund for Excellent Young Scholars (21822806).
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Z.J. Qin, Y. Chen, and S.Q. Yu performed experiments and data analysis. Z.J. Qin and J.W. Zhou wrote the manuscript and conceived the study. J. Chen and J.W. Zhou coordinated the project. All authors read and approved the final manuscript.
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Qin, Z., Chen, Y., Yu, S. et al. Engineering Gluconobacter cerinus CGMCC 1.110 for direct 2-keto-L-gulonic acid production. Appl Microbiol Biotechnol 107, 153–162 (2023). https://doi.org/10.1007/s00253-022-12310-5
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DOI: https://doi.org/10.1007/s00253-022-12310-5