Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli
Abstract Background Methanol has attracted increased attention as a non-food alternative carbon source to sugar for biological production of chemicals and fuels. Moreover, the high degree of reduction of methanol offers some advantages in increasing the production yields of NAD(P)H-dependent metabol...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2019-01-01
|
| Series: | Biotechnology for Biofuels |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s13068-019-1356-4 |
| _version_ | 1828811851687788544 |
|---|---|
| author | Xin Wang Xuelin Wang Xiaolu Lu Chen Ma Kequan Chen Pingkai Ouyang |
| author_facet | Xin Wang Xuelin Wang Xiaolu Lu Chen Ma Kequan Chen Pingkai Ouyang |
| author_sort | Xin Wang |
| collection | DOAJ |
| description | Abstract Background Methanol has attracted increased attention as a non-food alternative carbon source to sugar for biological production of chemicals and fuels. Moreover, the high degree of reduction of methanol offers some advantages in increasing the production yields of NAD(P)H-dependent metabolites. Here, we demonstrate an example of methanol bioconversion with the aim of improving production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli. Results A synthetic methylotrophic E. coli was engineered with a nicotinamide adenine dinucleotide (NAD+)-dependent methanol dehydrogenase (MDH) and ribulose monophosphate (RuMP) pathway. Regarding the limited MDH activity, the role of activator proteins in vivo was investigated, and the NudF protein was identified capable of improving MDH activity and triggering increased methanol metabolism. Using 13C-methanol-labeling experiments, we confirmed methanol assimilation in the methylotrophic E. coli. A cycling RuMP pathway for methanol assimilation was also demonstrated by detecting multiple labeled carbons for several compounds. Finally, using the NAD(P)H-dependent metabolite lysine as a test, the potential of methanol bioconversion to generate value-added metabolites was determined. To further characterize the benefit of methanol as the carbon source, extra NADH from methanol oxidation was engineered to generate NADPH to improve lysine biosynthesis by expression of the POS5 gene from Saccharomyces cerevisiae, which resulted in a twofold improvement of lysine production. Moreover, this new sink further pulled upstream methanol utilization. Conclusion Through engineering methanol metabolism, lysine biosynthesis, and NADPH regeneration pathway from NADH, the bioconversion of methanol to improve chemical synthesis was successfully achieved in methylotrophic E. coli. |
| first_indexed | 2024-12-12T09:34:32Z |
| format | Article |
| id | doaj.art-eb43bd51ec984e0d918057aa8418236d |
| institution | Directory Open Access Journal |
| issn | 1754-6834 |
| language | English |
| last_indexed | 2024-12-12T09:34:32Z |
| publishDate | 2019-01-01 |
| publisher | BMC |
| record_format | Article |
| series | Biotechnology for Biofuels |
| spelling | doaj.art-eb43bd51ec984e0d918057aa8418236d2022-12-22T00:28:46ZengBMCBiotechnology for Biofuels1754-68342019-01-0112111110.1186/s13068-019-1356-4Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coliXin Wang0Xuelin Wang1Xiaolu Lu2Chen Ma3Kequan Chen4Pingkai Ouyang5State Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech UniversityState Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech UniversityState Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech UniversityState Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech UniversityState Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech UniversityState Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech UniversityAbstract Background Methanol has attracted increased attention as a non-food alternative carbon source to sugar for biological production of chemicals and fuels. Moreover, the high degree of reduction of methanol offers some advantages in increasing the production yields of NAD(P)H-dependent metabolites. Here, we demonstrate an example of methanol bioconversion with the aim of improving production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli. Results A synthetic methylotrophic E. coli was engineered with a nicotinamide adenine dinucleotide (NAD+)-dependent methanol dehydrogenase (MDH) and ribulose monophosphate (RuMP) pathway. Regarding the limited MDH activity, the role of activator proteins in vivo was investigated, and the NudF protein was identified capable of improving MDH activity and triggering increased methanol metabolism. Using 13C-methanol-labeling experiments, we confirmed methanol assimilation in the methylotrophic E. coli. A cycling RuMP pathway for methanol assimilation was also demonstrated by detecting multiple labeled carbons for several compounds. Finally, using the NAD(P)H-dependent metabolite lysine as a test, the potential of methanol bioconversion to generate value-added metabolites was determined. To further characterize the benefit of methanol as the carbon source, extra NADH from methanol oxidation was engineered to generate NADPH to improve lysine biosynthesis by expression of the POS5 gene from Saccharomyces cerevisiae, which resulted in a twofold improvement of lysine production. Moreover, this new sink further pulled upstream methanol utilization. Conclusion Through engineering methanol metabolism, lysine biosynthesis, and NADPH regeneration pathway from NADH, the bioconversion of methanol to improve chemical synthesis was successfully achieved in methylotrophic E. coli.http://link.springer.com/article/10.1186/s13068-019-1356-4MethanolSynthetic methylotrophic E. coliCofactor regenerationNAD(P)H-dependent chemicals |
| spellingShingle | Xin Wang Xuelin Wang Xiaolu Lu Chen Ma Kequan Chen Pingkai Ouyang Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli Biotechnology for Biofuels Methanol Synthetic methylotrophic E. coli Cofactor regeneration NAD(P)H-dependent chemicals |
| title | Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli |
| title_full | Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli |
| title_fullStr | Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli |
| title_full_unstemmed | Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli |
| title_short | Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli |
| title_sort | methanol fermentation increases the production of nad p h dependent chemicals in synthetic methylotrophic escherichia coli |
| topic | Methanol Synthetic methylotrophic E. coli Cofactor regeneration NAD(P)H-dependent chemicals |
| url | http://link.springer.com/article/10.1186/s13068-019-1356-4 |
| work_keys_str_mv | AT xinwang methanolfermentationincreasestheproductionofnadphdependentchemicalsinsyntheticmethylotrophicescherichiacoli AT xuelinwang methanolfermentationincreasestheproductionofnadphdependentchemicalsinsyntheticmethylotrophicescherichiacoli AT xiaolulu methanolfermentationincreasestheproductionofnadphdependentchemicalsinsyntheticmethylotrophicescherichiacoli AT chenma methanolfermentationincreasestheproductionofnadphdependentchemicalsinsyntheticmethylotrophicescherichiacoli AT kequanchen methanolfermentationincreasestheproductionofnadphdependentchemicalsinsyntheticmethylotrophicescherichiacoli AT pingkaiouyang methanolfermentationincreasestheproductionofnadphdependentchemicalsinsyntheticmethylotrophicescherichiacoli |