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Abstract
BackgroundThere is a strong interest in using photosynthetic cyanobacteria as production hosts for biofuels and chemicals. Recent work has shown the benefit of pathway engineering, enzyme tolerance, and co-factor usage for improving yields of fermentation products.ResultsAn n-butanol pathway was inserted into a Synechocystis mutant deficient in polyhydroxybutyrate synthesis. We found that nitrogen starvation increased specific butanol productivity up to threefold, but cessation of cell growth limited total n-butanol titers. Metabolite profiling showed that acetyl-CoA increased twofold during nitrogen starvation. Introduction of a phosphoketolase increased acetyl-CoA levels sixfold at nitrogen replete conditions and increased butanol titers from 22 to 37 mg/L at day 8. Flux balance analysis of photoautotrophic metabolism showed that a Calvin–Benson–Bassham-Phosphoketolase pathway had higher theoretical butanol productivity than CBB-Embden–Meyerhof–Parnas and a reduced butanol ATP demand.ConclusionThese results demonstrate that phosphoketolase overexpression and modulation of nitrogen levels are two attractive routes toward increased production of acetyl-CoA derived products in cyanobacteria and could be implemented with complementary metabolic engineering strategies.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0355-9) contains supplementary material, which is available to authorized users.
Highlights
There is a strong interest in using photosynthetic cyanobacteria as production hosts for biofuels and chemicals
The results show the utility of phosphoketolase in re-directing autotrophic metabolism, which could be used in addition to other metabolic engineering strategies
Expression of butanol‐production pathway in nitrogen replete and deplete conditions The n-butanol fermentation pathway of Clostridia proceeds from acetyl-CoA in six steps and has been adapted to other bacterial hosts [12, 13] as well as yeast [14, 15]
Summary
There is a strong interest in using photosynthetic cyanobacteria as production hosts for biofuels and chemicals. Recent work has shown the benefit of pathway engineering, enzyme tolerance, and co-factor usage for improving yields of fermentation products. Photosynthetic microbes are promising chemical and biofuel hosts as they circumvent the use of plant-based feedstocks and eliminate a large cost for currentgeneration biofuels. Cyanobacteria in particular are attractive due to extensive genetic and biochemical characterization, ease of genetic modification, and diverse ecology and habitat. Recent work has aimed to increase metabolic flux toward biofuel production in cyanobacteria. Nitrogen content of the culture media can induce drastic changes to the central carbon metabolism of nondiazotrophic cyanobacteria such as Synechocystis. Carbon is directed to storage compounds such as glycogen and polyhydroxybutyrate (PHB)
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