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Modular design of metabolic network for robust production of n-butanol from galactose-glucose mixtures.

Lim HG, Lim JH, Jung GY - Biotechnol Biofuels (2015)

Bottom Line: Specifically, the engineered strain showed dramatically increased n-butanol production (3.3-fold increased to 6.2 g/L after 48-h fermentation) compared to the parental strain (1.9 g/L) in galactose-supplemented medium.Collectively, modular pathway engineering of metabolic network can be an effective approach in strain development for optimal biofuel production with cost-effective fermentable sugars.Moreover, robust production of n-butanol with sugar mixtures with variable composition would facilitate the economic feasibility of the microbial process using a mixture of sugars from cheap biomass in the near future.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical Engineering, Pohang University of Science and Technology, 77 Cheongam-Ro, Pohang, 37673 Gyeongbuk Korea.

ABSTRACT

Background: Refactoring microorganisms for efficient production of advanced biofuel such as n-butanol from a mixture of sugars in the cheap feedstock is a prerequisite to achieve economic feasibility in biorefinery. However, production of biofuel from inedible and cheap feedstock is highly challenging due to the slower utilization of biomass-driven sugars, arising from complex assimilation pathway, difficulties in amplification of biosynthetic pathways for heterologous metabolite, and redox imbalance caused by consuming intracellular reducing power to produce quite reduced biofuel. Even with these problems, the microorganisms should show robust production of biofuel to obtain industrial feasibility. Thus, refactoring microorganisms for efficient conversion is highly desirable in biofuel production.

Results: In this study, we engineered robust Escherichia coli to accomplish high production of n-butanol from galactose-glucose mixtures via the design of modular pathway, an efficient and systematic way, to reconstruct the entire metabolic pathway with many target genes. Three modular pathways designed using the predictable genetic elements were assembled for efficient galactose utilization, n-butanol production, and redox re-balancing to robustly produce n-butanol from a sugar mixture of galactose and glucose. Specifically, the engineered strain showed dramatically increased n-butanol production (3.3-fold increased to 6.2 g/L after 48-h fermentation) compared to the parental strain (1.9 g/L) in galactose-supplemented medium. Moreover, fermentation with mixtures of galactose and glucose at various ratios from 2:1 to 1:2 confirmed that our engineered strain was able to robustly produce n-butanol regardless of sugar composition with simultaneous utilization of galactose and glucose.

Conclusions: Collectively, modular pathway engineering of metabolic network can be an effective approach in strain development for optimal biofuel production with cost-effective fermentable sugars. To the best of our knowledge, this study demonstrated the first and highest n-butanol production from galactose in E. coli. Moreover, robust production of n-butanol with sugar mixtures with variable composition would facilitate the economic feasibility of the microbial process using a mixture of sugars from cheap biomass in the near future.

No MeSH data available.


Related in: MedlinePlus

Time-course of fermentation profiles of the parental strain (JHL80, a through c) and redox optimized strain (GAL_083, d through f) in mixtures of galactose and glucose supplemented medium for 48 h. The pH was adjusted to around 7.2 at 6-h intervals. The ratio of galactose to glucose was varied from 2:1 (a, d), to 1:1 (b, e), to 1:2 (c, f). The GAL_083 strain showed the ability to utilize galactose and glucose simultaneously. The lefty-offset and righty-axis represent sugar and metabolite (n-butanol, ethanol, and butyrate) concentrations (g/L), respectively. The lefty-axis represents OD600. The x-axis represents time in culture (h). The error bars indicate standard deviations of measurements from two independent cultures. Symbols: open circle OD600; open rectangle galactose; closed rectangle glucose; closed up trianglen-butanol; closed diamond ethanol; closed down triangle butyrate
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Fig5: Time-course of fermentation profiles of the parental strain (JHL80, a through c) and redox optimized strain (GAL_083, d through f) in mixtures of galactose and glucose supplemented medium for 48 h. The pH was adjusted to around 7.2 at 6-h intervals. The ratio of galactose to glucose was varied from 2:1 (a, d), to 1:1 (b, e), to 1:2 (c, f). The GAL_083 strain showed the ability to utilize galactose and glucose simultaneously. The lefty-offset and righty-axis represent sugar and metabolite (n-butanol, ethanol, and butyrate) concentrations (g/L), respectively. The lefty-axis represents OD600. The x-axis represents time in culture (h). The error bars indicate standard deviations of measurements from two independent cultures. Symbols: open circle OD600; open rectangle galactose; closed rectangle glucose; closed up trianglen-butanol; closed diamond ethanol; closed down triangle butyrate

Mentions: Under these conditions, the engineered strain utilized galactose and glucose simultaneously, while the parental strain showed no galactose consumption until glucose was depleted (Fig. 5). Moreover, n-butanol production by the engineered strain was consistently above 5 g/L regardless of the composition of the medium, while the yield of metabolites produced by the parental strain varied. Taken together, these results indicate that this strain would be promising as a platform strain for the optimal and robust production of valuable bio-chemicals from various galactose–glucose mixtures without regard to possible changes in the composition.Fig. 5


Modular design of metabolic network for robust production of n-butanol from galactose-glucose mixtures.

Lim HG, Lim JH, Jung GY - Biotechnol Biofuels (2015)

Time-course of fermentation profiles of the parental strain (JHL80, a through c) and redox optimized strain (GAL_083, d through f) in mixtures of galactose and glucose supplemented medium for 48 h. The pH was adjusted to around 7.2 at 6-h intervals. The ratio of galactose to glucose was varied from 2:1 (a, d), to 1:1 (b, e), to 1:2 (c, f). The GAL_083 strain showed the ability to utilize galactose and glucose simultaneously. The lefty-offset and righty-axis represent sugar and metabolite (n-butanol, ethanol, and butyrate) concentrations (g/L), respectively. The lefty-axis represents OD600. The x-axis represents time in culture (h). The error bars indicate standard deviations of measurements from two independent cultures. Symbols: open circle OD600; open rectangle galactose; closed rectangle glucose; closed up trianglen-butanol; closed diamond ethanol; closed down triangle butyrate
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4559943&req=5

Fig5: Time-course of fermentation profiles of the parental strain (JHL80, a through c) and redox optimized strain (GAL_083, d through f) in mixtures of galactose and glucose supplemented medium for 48 h. The pH was adjusted to around 7.2 at 6-h intervals. The ratio of galactose to glucose was varied from 2:1 (a, d), to 1:1 (b, e), to 1:2 (c, f). The GAL_083 strain showed the ability to utilize galactose and glucose simultaneously. The lefty-offset and righty-axis represent sugar and metabolite (n-butanol, ethanol, and butyrate) concentrations (g/L), respectively. The lefty-axis represents OD600. The x-axis represents time in culture (h). The error bars indicate standard deviations of measurements from two independent cultures. Symbols: open circle OD600; open rectangle galactose; closed rectangle glucose; closed up trianglen-butanol; closed diamond ethanol; closed down triangle butyrate
Mentions: Under these conditions, the engineered strain utilized galactose and glucose simultaneously, while the parental strain showed no galactose consumption until glucose was depleted (Fig. 5). Moreover, n-butanol production by the engineered strain was consistently above 5 g/L regardless of the composition of the medium, while the yield of metabolites produced by the parental strain varied. Taken together, these results indicate that this strain would be promising as a platform strain for the optimal and robust production of valuable bio-chemicals from various galactose–glucose mixtures without regard to possible changes in the composition.Fig. 5

Bottom Line: Specifically, the engineered strain showed dramatically increased n-butanol production (3.3-fold increased to 6.2 g/L after 48-h fermentation) compared to the parental strain (1.9 g/L) in galactose-supplemented medium.Collectively, modular pathway engineering of metabolic network can be an effective approach in strain development for optimal biofuel production with cost-effective fermentable sugars.Moreover, robust production of n-butanol with sugar mixtures with variable composition would facilitate the economic feasibility of the microbial process using a mixture of sugars from cheap biomass in the near future.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical Engineering, Pohang University of Science and Technology, 77 Cheongam-Ro, Pohang, 37673 Gyeongbuk Korea.

ABSTRACT

Background: Refactoring microorganisms for efficient production of advanced biofuel such as n-butanol from a mixture of sugars in the cheap feedstock is a prerequisite to achieve economic feasibility in biorefinery. However, production of biofuel from inedible and cheap feedstock is highly challenging due to the slower utilization of biomass-driven sugars, arising from complex assimilation pathway, difficulties in amplification of biosynthetic pathways for heterologous metabolite, and redox imbalance caused by consuming intracellular reducing power to produce quite reduced biofuel. Even with these problems, the microorganisms should show robust production of biofuel to obtain industrial feasibility. Thus, refactoring microorganisms for efficient conversion is highly desirable in biofuel production.

Results: In this study, we engineered robust Escherichia coli to accomplish high production of n-butanol from galactose-glucose mixtures via the design of modular pathway, an efficient and systematic way, to reconstruct the entire metabolic pathway with many target genes. Three modular pathways designed using the predictable genetic elements were assembled for efficient galactose utilization, n-butanol production, and redox re-balancing to robustly produce n-butanol from a sugar mixture of galactose and glucose. Specifically, the engineered strain showed dramatically increased n-butanol production (3.3-fold increased to 6.2 g/L after 48-h fermentation) compared to the parental strain (1.9 g/L) in galactose-supplemented medium. Moreover, fermentation with mixtures of galactose and glucose at various ratios from 2:1 to 1:2 confirmed that our engineered strain was able to robustly produce n-butanol regardless of sugar composition with simultaneous utilization of galactose and glucose.

Conclusions: Collectively, modular pathway engineering of metabolic network can be an effective approach in strain development for optimal biofuel production with cost-effective fermentable sugars. To the best of our knowledge, this study demonstrated the first and highest n-butanol production from galactose in E. coli. Moreover, robust production of n-butanol with sugar mixtures with variable composition would facilitate the economic feasibility of the microbial process using a mixture of sugars from cheap biomass in the near future.

No MeSH data available.


Related in: MedlinePlus