Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production
Abstract 5-aminolevulinic acid (ALA) has broad potential applications in the medical, agricultural and food industries. Several strategies have been implemented successfully to try to improve ALA synthesis. Nonetheless, the low yield has got in the way of large-scale bio-manufacture of 5-ALA. In thi...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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SpringerOpen
2021-12-01
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| Series: | AMB Express |
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| Online Access: | https://doi.org/10.1186/s13568-021-01335-0 |
| _version_ | 1819173911846191104 |
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| author | Fanglan Ge Xiaokun Li Qingrong Ge Di Zhu Wei Li Fenghui Shi Hongjin Chen |
| author_facet | Fanglan Ge Xiaokun Li Qingrong Ge Di Zhu Wei Li Fenghui Shi Hongjin Chen |
| author_sort | Fanglan Ge |
| collection | DOAJ |
| description | Abstract 5-aminolevulinic acid (ALA) has broad potential applications in the medical, agricultural and food industries. Several strategies have been implemented successfully to try to improve ALA synthesis. Nonetheless, the low yield has got in the way of large-scale bio-manufacture of 5-ALA. In this study, we explored strain engineering strategies for high‐level 5‐ALA production in Corynebacterium glutamicum F343 using the C4 pathway. Initially, the glutamate dehydrogenase-encoding gene gdhA was deleted to reduce glutamate yield. Then the C4 pathway was introduced in the gdhA mutant strain F2-A (∆gdhA + hemA), resulting in a 5-ALA yield of up to 3.2 g/L. Furthermore, the accumulations of downstream metabolites such as heme, porphobilinogen, and protoporphyrin IX, were decreased. After evaluating the mechanisms of this synthetic pathway by RNA-Seq, the results showed that genes involved in both the C5 pathway and heme pathways were down-regulated in strain F2-A (∆gdhA + hemA). Interestingly, upstream genes of succinyl-CoA in the tricarboxylic acid (TCA) cycle, such as icd, lpdA, were up-regulated, while its downstream genes, including sucC, sucD, sdhB, sdhA, sdhCD, were down-regulated. These changes amplify the sources of succinyl-CoA and reduce its expenditure, before pulling the carbon flux to produce 5-ALA. Furthermore, the down-regulation of most genes of the heme pathway could reduce the drainage of 5‐ALA, which further enhance its accumulation. To alleviate competition between glyoxylate and the TCA cycle, the isocitrate dehydrogenase-encoding gene aceA was also knocked out, resulting in 3.86 g/L of 5‐ALA. Finally, the fermentation conditions were optimized, resulting in a maximum 5-ALA yield of 5.6 g/L. Overall, the blocking of the glutamate synthesis pathway could be a powerful strategy to re-allocate the carbon flux to produce 5-ALA. It could also enable the efficient synthesis of other TCA derivatives in C. glutamicum. |
| first_indexed | 2024-12-22T20:30:36Z |
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| id | doaj.art-fe819af97371469f8e26ac5ecd157ef4 |
| institution | Directory Open Access Journal |
| issn | 2191-0855 |
| language | English |
| last_indexed | 2024-12-22T20:30:36Z |
| publishDate | 2021-12-01 |
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| series | AMB Express |
| spelling | doaj.art-fe819af97371469f8e26ac5ecd157ef42022-12-21T18:13:36ZengSpringerOpenAMB Express2191-08552021-12-0111111210.1186/s13568-021-01335-0Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid productionFanglan Ge0Xiaokun Li1Qingrong Ge2Di Zhu3Wei Li4Fenghui Shi5Hongjin Chen6College of Life Sciences, Sichuan Normal UniversityCollege of Life Sciences, Sichuan Normal UniversityCollege of Life Sciences, Sichuan Normal UniversityCollege of Life Sciences, Sichuan Normal UniversityCollege of Life Sciences, Sichuan Normal UniversityCollege of Life Sciences, Sichuan Normal UniversityCollege of Life Sciences, Sichuan Normal UniversityAbstract 5-aminolevulinic acid (ALA) has broad potential applications in the medical, agricultural and food industries. Several strategies have been implemented successfully to try to improve ALA synthesis. Nonetheless, the low yield has got in the way of large-scale bio-manufacture of 5-ALA. In this study, we explored strain engineering strategies for high‐level 5‐ALA production in Corynebacterium glutamicum F343 using the C4 pathway. Initially, the glutamate dehydrogenase-encoding gene gdhA was deleted to reduce glutamate yield. Then the C4 pathway was introduced in the gdhA mutant strain F2-A (∆gdhA + hemA), resulting in a 5-ALA yield of up to 3.2 g/L. Furthermore, the accumulations of downstream metabolites such as heme, porphobilinogen, and protoporphyrin IX, were decreased. After evaluating the mechanisms of this synthetic pathway by RNA-Seq, the results showed that genes involved in both the C5 pathway and heme pathways were down-regulated in strain F2-A (∆gdhA + hemA). Interestingly, upstream genes of succinyl-CoA in the tricarboxylic acid (TCA) cycle, such as icd, lpdA, were up-regulated, while its downstream genes, including sucC, sucD, sdhB, sdhA, sdhCD, were down-regulated. These changes amplify the sources of succinyl-CoA and reduce its expenditure, before pulling the carbon flux to produce 5-ALA. Furthermore, the down-regulation of most genes of the heme pathway could reduce the drainage of 5‐ALA, which further enhance its accumulation. To alleviate competition between glyoxylate and the TCA cycle, the isocitrate dehydrogenase-encoding gene aceA was also knocked out, resulting in 3.86 g/L of 5‐ALA. Finally, the fermentation conditions were optimized, resulting in a maximum 5-ALA yield of 5.6 g/L. Overall, the blocking of the glutamate synthesis pathway could be a powerful strategy to re-allocate the carbon flux to produce 5-ALA. It could also enable the efficient synthesis of other TCA derivatives in C. glutamicum.https://doi.org/10.1186/s13568-021-01335-05-aminolevulinic acidCorynebacteriumglutamicumHeme biosynthesisPathways engineeringGlutamateRNA-Seq analysis |
| spellingShingle | Fanglan Ge Xiaokun Li Qingrong Ge Di Zhu Wei Li Fenghui Shi Hongjin Chen Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production AMB Express 5-aminolevulinic acid Corynebacteriumglutamicum Heme biosynthesis Pathways engineering Glutamate RNA-Seq analysis |
| title | Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production |
| title_full | Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production |
| title_fullStr | Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production |
| title_full_unstemmed | Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production |
| title_short | Modular control of multiple pathways of Corynebacterium glutamicum for 5-aminolevulinic acid production |
| title_sort | modular control of multiple pathways of corynebacterium glutamicum for 5 aminolevulinic acid production |
| topic | 5-aminolevulinic acid Corynebacteriumglutamicum Heme biosynthesis Pathways engineering Glutamate RNA-Seq analysis |
| url | https://doi.org/10.1186/s13568-021-01335-0 |
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