Carbon dioxide fixation by Calvin-Cycle enzymes improves ethanol yield in yeast

Biotechnology for Biofuels

Víctor Guadalupe-Medina¹  H Wouter Wisselink¹  Marijke AH Luttik¹  Erik de Hulster¹  Jean-Marc Daran¹  Jack T Pronk¹  Antonius JA van Maris¹ 

[1] Kluyver Centre for Genomics of Industrial Fermentation, P.O. Box 5057, 2600, GA Delft, The Netherlands

关键词: Bioethanol;    Glycerol;    Saccharomyces cerevisiae;    Carbon dioxide fixation;    NADH re-oxidation;    Phosphoribulokinase;    Ribulose-1,5-bisphosphate carboxylase;    Rubisco;    Synthetic biology;    Metabolic engineering;   

Others  :  797922
DOI  :  10.1186/1754-6834-6-125

 received in 2013-06-20, accepted in 2013-08-27,  发布年份 2013

【 摘 要 】

Background

Redox-cofactor balancing constrains product yields in anaerobic fermentation processes. This challenge is exemplified by the formation of glycerol as major by-product in yeast-based bioethanol production, which is a direct consequence of the need to reoxidize excess NADH and causes a loss of conversion efficiency. Enabling the use of CO₂ as electron acceptor for NADH oxidation in heterotrophic microorganisms would increase product yields in industrial biotechnology.

Results

A hitherto unexplored strategy to address this redox challenge is the functional expression in yeast of enzymes from autotrophs, thereby enabling the use of CO₂ as electron acceptor for NADH reoxidation. Functional expression of the Calvin cycle enzymes phosphoribulokinase (PRK) and ribulose-1,5-bisphosphate carboxylase (Rubisco) in Saccharomyces cerevisiae led to a 90% reduction of the by-product glycerol and a 10% increase in ethanol production in sugar-limited chemostat cultures on a mixture of glucose and galactose. Co-expression of the Escherichia coli chaperones GroEL and GroES was key to successful expression of CbbM, a form-II Rubisco from the chemolithoautotrophic bacterium Thiobacillus denitrificans in yeast.

Conclusions

Our results demonstrate functional expression of Rubisco in a heterotrophic eukaryote and demonstrate how incorporation of CO₂ as a co-substrate in metabolic engineering of heterotrophic industrial microorganisms can be used to improve product yields. Rapid advances in molecular biology should allow for rapid insertion of this 4-gene expression cassette in industrial yeast strains to improve production, not only of 1st and 2nd generation ethanol production, but also of other renewable fuels or chemicals.

【 授权许可】

   
2013 Guadalupe-Medina et al.; licensee BioMed Central Ltd.

【 图 表 】

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