Abstract
BackgroundPropionate is widely used as an important preservative and important chemical intermediate for synthesis of cellulose fibers, herbicides, perfumes and pharmaceuticals. Biosynthetic propionate has mainly been produced by Propionibacterium, which has various limitations for industrial application.ResultsIn this study, we engineered E. coli by combining reduced TCA cycle with the native sleeping beauty mutase (Sbm) cycle to construct a redox balanced and energy viable fermentation pathway for anaerobic propionate production. As the cryptic Sbm operon was over-expressed in E. coli MG1655, propionate titer reached 0.24 g/L. To increase precursor supply for the Sbm cycle, genetic modification was made to convert mixed fermentation products to succinate, which slightly increased propionate production. For optimal expression of Sbm operon, different types of promoters were examined. A strong constitutive promoter Pbba led to the highest titer of 2.34 g/L. Methylmalonyl CoA mutase from Methylobacterium extorquens AM1 was added to strain T110(pbba-Sbm) to enhance this rate limiting step. With optimized expression of this additional Methylmalonyl CoA mutase, the highest production strain was obtained with a titer of 4.95 g/L and a yield of 0.49 mol/mol glucose.ConclusionsWith various metabolic engineering strategies, the propionate titer from fermentation achieved 4.95 g/L. This is the reported highest anaerobic production of propionate by heterologous host. Due to host advantages, such as non-strict anaerobic condition, mature engineering and fermentation techniques, and low cost minimal media, our work has built the basis for industrial propionate production with E. coli chassis.
Highlights
Propionate is widely used as an important preservative and important chemical intermediate for synthesis of cellulose fibers, herbicides, perfumes and pharmaceuticals
By studying reduced TCA cycle and sleeping beauty mutase (Sbm) cycle, we designed a novel anaerobic pathway from glucose to propionate as illustrated in Fig. 1, indicated by bold arrows. This pathway goes through reduced TCA cycle from Phosphoenolpyruvate (PEP) to succinate, which is converted to propionate by Sbm cycle
This pathway goes through reduced TCA cycle from PEP to succinate, which is converted to propionate by Sbm cycle
Summary
Propionate is widely used as an important preservative and important chemical intermediate for synthesis of cellulose fibers, herbicides, perfumes and pharmaceuticals. Biosynthetic propionate has mainly been produced by Propionibacterium, which has various limitations for industrial application. Propionate is widely used as an important preservative in animal feed and human foods, which is a chemical intermediate for synthesis of cellulose fibers, herbicides, perfumes and pharmaceuticals [1, 2]. Fermentation by propionibacteria has various limitations, such as nitrogen flux for maintaining anaerobic condition [15], slow growth,costly complex culture media and lack of metabolic engineering tools for strain improvement [17], which make this technology not economically applicable
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