Engineered Non-Oxidative Glycolysis Pathway For Acetone And Isopropanol Production In Escherichia Coli

Acetone and isopropanol are important chemical raw material and organic solvent.Considering the energy and environmental problems,acetone and isopropanol production from renewable bioresources is an attractive alternative.In this study,we introduced the acetone and isopropanol synthetic pathway into Escherichia coli.In shake flask cultures,the titer of strain JM109 harboring the acetone and isopropanol synthesis pathway were 38 m M and 35 m M respectively after 22 h cultivation at a yeast extract concentration of 2 g/L.In E.coli,acetyl-Co A is mainly produced by decarboxylation of pyruvate whereby one carbon equivalent is lost as CO₂.Consequently,the maximal theoretical molar yield of acetone and isopropanol from glucose is only 1?two acetyl-Co A to one acetone?.To find new metabolic engineering strategies to improve acetone and isopropanol yield in Escherichia coli,we conducted flux balance analysis.Flux balance analysis results from a modified E.coli genome scale metabolic network suggested that the introduction of a non-oxidative glycolysis pathway can improve the theoretical acetone and isopropanol yield from 1 to 1.5 mol acetone/mol glucose and 1to 1.33 mol isopropanol/mol glucose.By inserting the phophoketolase gene?fxpk?from Bifidobacterium adolescentis into the genome,we constructed a non-oxidative glycolysis pathway in E.coli.The resulting strain NOG produced 47 m M acetone and 49 m M isopropanol from glucose under aerobic conditions in shake flasks.The mole yield of acetone was improved from 38% to 47% and the mole yield of isopropanol was improved from 35% to 40%.To improve the flux of the non-oxidative glycolysis pathway,we overexpressed the transaldolase?tal A?and transketolase?tkt A?in the strain NOG genome by inserting the promoter J23100.The mole yield of isopropanol from glucose improved from 40% to 49%.To reduce the main by-product acetate concentration,we overexpress the acetyl-Co A synthetase in the NOG genome by inserting the promoter J23100.The peptidyl-lysine acetyltransferase gene?pka?was deleted from the genome to reduce the acetylation effect.The mole yield of isopropanol from glucose improved from40% to 47%.