Abstract
l-cysteine, a valuable sulfur-containing amino acid, has been widely used in food, agriculture, and pharmaceutical industries. Due to the toxicity and complex regulation of l-cysteine, no efficient cell factory has yet been achieved for l-cysteine industrial production. In this study, the food-grade microorganism Corynebacterium glutamicum was engineered for l-cysteine production. Through deletion of the l-cysteine desulfhydrases (CD) and overexpression of the native serine acetyltransferase (CysE), the initial l-cysteine-producing strain CYS-2 was constructed to produce 58.2 ± 5.1 mg/L of l-cysteine. Subsequently, several metabolic engineering strategies were performed to further promote l-cysteine biosynthesis, including using strong promoter tac-M to enhance expression intensity of CysE, investigating the best candidate among several heterogeneous feedback-insensitive CysEs for l-cysteine biosynthesis, overexpressing l-cysteine synthase (CysK) to drive more metabolic flux, evaluating the efflux capacity of several heterogeneous l-cysteine transporters, engineering l-serine biosynthesis module to increase the precursor l-serine level and using thiosulfate as the sulfur source. Finally, the l-cysteine concentration of the engineered strain CYS-19 could produce 947.9 ± 46.5 mg/L with addition of 6 g/L Na2S2O3, approximately 14.1-fold higher than that of the initial strain CYS-2, which was the highest titer of l-cysteine ever reported in C. glutamicum. These results indicated that C. glutamicum was a promising platform for l-cysteine production.
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Acknowledgements
We are grateful to Prof. Masayuki Inui (Research Institute of Innovative Technology for the Earth, Japan) for generously providing the plasmids.
Funding
This study was supported by the National Natural Science Foundation of China (No. 31500044), the Natural Science Foundation of Tianjin (No. 17JCQNJC09600, No. 17JCYBJC24000), the Foundation of Hebei Educational Committee (ZD2017047), the Tianjin Science and Technology Project (15PTCYSY00020), and the Key Projects in the Tianjin Science and Technology Pillar Program (14ZCZDSY00058), as well as “Hundred Talents Program” of the Chinese Academy of Sciences.
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Wei, L., Wang, H., Xu, N. et al. Metabolic engineering of Corynebacterium glutamicum for l-cysteine production. Appl Microbiol Biotechnol 103, 1325–1338 (2019). https://doi.org/10.1007/s00253-018-9547-7
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DOI: https://doi.org/10.1007/s00253-018-9547-7