Enhanced 2,3-butanediol production from biodiesel-derived glycerol by engineering of cofactor regeneration and manipulating carbon flux in Bacillus amyloliquefaciens.

Taowei Yang,Zhenghong Xu,Meijuan Xu,Xian Zhang,Zhiming Rao,Shang-Tian Yang

doi:10.1186/s12934-015-0317-2

Taowei Yang, Zhenghong Xu + Show 4 more

Open Access

https://doi.org/10.1186/s12934-015-0317-2

Copy DOI

Abstract

BackgroundBacillus amyloliquefaciens B10-127 exhibited an excellent ability for industrial-scale microbial fermentation of 2,3-butanediol (2,3-BD) from biodiesel-derived glycerol. However, the accumulation of by-products (acetoin, acetoin, lactate and succinate) and the 2,3-BD yield remains prohibitively low for commercial production.ResultsSeveral strategies were developed to manipulate the carbon flux to 2,3-BD branch in a designed B. amyloliquefaciens. Firstly, extra copies of NADH/NAD+ regeneration system were introduced into B. amyloliquefaciens by co-overproduction of glycerol dehydrogenase and acetoin reductase, which resulting in improvement of 2,3-BD production and suppression of by-products accumulation. Subsequently, the transcriptional regulator ALsR under the control of a moderate promoter PbdhA was introduced into B. amyloliquefaciens, which increased carbon flux to 2,3-BD branch. Finally, a three-stage dissolved oxygen control strategy were proposed based on analysis of the characteristic of 2,3-BD fermentation, and a two-stage pH control strategy were proposed based on different pH preferences of ACR for reduction and oxidation. Following these strategies, a high titer (102.3 g/L), yield (0.44 g/g), and productivity (1.16 g/L/h) of 2,3-BD were achieved.ConclusionsTo our knowledge, this is the highest reported 2,3-BD production using biodiesel-derived glycerol as substrate, and this designed B. amyloliquefaciens should be an excellent candidate for producing 2,3-BD on an industrial scale.

Highlights

Introduction of extra copies ofDhaD/acetoin reductase (ACR) enzymes into B. amyloliquefaciens and their effects on 2,3‐BD production The NAD+-dependent DhaD oxidizes glycerol to dihydroxyacetone, with concomitant reduction of NAD+ to NADH
Extra copies of NADH/NAD+ regeneration system were introduced into B. amyloliquefaciens by co-overproduction of glycerol dehydrogenase and acetoin reductase
A three-stage dissolved oxygen control strategy were proposed based on analysis of the characteristic of 2,3-BD fermentation, and a two-stage pH control strategy were proposed based on different pH preferences of ACR for reduction and oxidation this coenzyme is preferentially used in 2,3-BD synthesis

Summary

Results

Several strategies were developed to manipulate the carbon flux to 2,3-BD branch in a designed B. amyloliquefaciens. The transcriptional regulator ALsR under the control of a moderate promoter PbdhA was introduced into B. amyloliquefaciens, which increased carbon flux to 2,3-BD branch. A three-stage dissolved oxygen control strategy were proposed based on analysis of the characteristic of 2,3-BD fermentation, and a two-stage pH control strategy were proposed based on different pH preferences of ACR for reduction and oxidation. Following these strategies, a high titer (102.3 g/L), yield (0.44 g/g), and productivity (1.16 g/L/h) of 2,3-BD were achieved

Conclusions

Results and discussion

Conclusion

Methods