Abstract
Spermidine is an important polyamine that can be used for the synthesis of various bioactive compounds in the food and pharmaceutical fields. In this study, a novel efficient whole-cell biocatalytic method with an NADPH self-sufficient cycle for spermidine biosynthesis was designed and constructed by co-expressing homoserine dehydrogenase (HSD), carboxyspermidine dehydrogenase (CASDH), and carboxyspermidine decarboxylase (CASDC). First, the enzyme–substrate coupled cofactor regeneration system from co-expression of NADP+-dependent ScHSD and NADPH-dependent AfCASDH exactly provides an efficient method for cofactor cycling. Second, we identified and characterized a putative CASDC with high decarboxylase activity from Butyrivibrio crossotus DSM 2876; it showed an optimum temperature of 35 °C and an optimum pH of 7.0, which make it better suited for the designed synthetic route. Subsequently, the protein expression level of each enzyme was optimized through the variation of the gene copy number, and a whole-cell catalyst with high catalytic efficiency was constructed successfully. Finally, a yield of 28.6 mM of spermidine was produced in a 1-L scale of E. coli whole-cell catalytic system with a 95.3% molar conversion rate after optimization of temperature, the ratio of catalyst-to-substrate, and the amount of NADP+, and a productivity of 0.17 g·L−1·h−1 was achieved. In summary, this novel pathway of constructing a whole-cell catalytic system from L-homoserine and putrescine could provide a green alternative method for the efficient synthesis of spermidine.
Key points
• A novel pathway for spermidine biosynthesis was developed in Escherichia coli.
• The enzyme–substrate coupled system provides an NADPH self-sufficient cycle.
• Spermidine with 28.6 mM was obtained using an optimized whole-cell system.
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Funding
This study was funded by the National Key Scientific Instrument and Equipment Development Project of China (2013YQ17052504), the Program for Changjiang Scholars and Innovative Research Team in the University of Ministry of Education of China (IRT_15R55), the seventh group of Hundred-Talent Program of Shanxi Province (2015), The Key Project of Research and Development Plan of Shaanxi (2017ZDCXL-SF-01–02-01), the Natural Sciences Foundation of Jiangsu Province (BK20210471), and the Natural Sciences Foundation of Guangdong Province (2021A1515110263).
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X. L. conceived and designed the research, conducted experimental work, interpreted the data, finalized conclusions, and wrote the manuscript. H. D. and T. F. analyzed the data and helped perform the analysis with constructive discussions. Y. B. and X. Z. contributed in funding acquisition. Y. C. conceived and designed the research, finalized conclusions, and wrote the manuscript. All authors contributed to the article and approved the submitted version.
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Liang, X., Deng, H., Bai, Y. et al. Highly efficient biosynthesis of spermidine from L-homoserine and putrescine using an engineered Escherichia coli with NADPH self-sufficient system. Appl Microbiol Biotechnol 106, 5479–5493 (2022). https://doi.org/10.1007/s00253-022-12110-x
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DOI: https://doi.org/10.1007/s00253-022-12110-x