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Quantitative target analysis and kinetic profiling of acyl-CoAs reveal the rate-limiting step in cyanobacterial 1-butanol production
Cyanobacterial 1-butanol production is an important model system for direct conversion of CO(2) to fuels and chemicals. Metabolically-engineered cyanobacteria introduced with a heterologous Coenzyme A (CoA)-dependent pathway modified from Clostridium species can convert atmospheric CO(2) into 1-buta...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Springer US
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4700068/ https://www.ncbi.nlm.nih.gov/pubmed/26766939 http://dx.doi.org/10.1007/s11306-015-0940-2 |
Sumario: | Cyanobacterial 1-butanol production is an important model system for direct conversion of CO(2) to fuels and chemicals. Metabolically-engineered cyanobacteria introduced with a heterologous Coenzyme A (CoA)-dependent pathway modified from Clostridium species can convert atmospheric CO(2) into 1-butanol. Efforts to optimize the 1-butanol pathway in Synechococcus elongatus PCC 7942 have focused on the improvement of the CoA-dependent pathway thus, probing the in vivo metabolic state of the CoA-dependent pathway is essential for identifying its limiting steps. In this study, we performed quantitative target analysis and kinetic profiling of acyl-CoAs in the CoA-dependent pathway by reversed phase ion-pair liquid chromatography-triple quadrupole mass spectrometry. Using (13)C-labelled cyanobacterial cell extract as internal standard, measurement of the intracellular concentration of acyl-CoAs revealed that the reductive reaction of butanoyl-CoA to butanal is a possible rate-limiting step. In addition, improvement of the butanoyl-CoA to butanal reaction resulted in an increased rate of acetyl-CoA synthesis by possibly compensating for the limitation of free CoA species. We inferred that the efficient recycling of free CoA played a key role in enhancing the conversion of pyruvate to acetyl-CoA. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11306-015-0940-2) contains supplementary material, which is available to authorized users. |
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