title: MEP pathway-mediated isopentenol production in metabolically engineered Escherichia coli.
abstract: BACKGROUND
  Isopentenols, such as prenol and isoprenol, are promising advanced biofuels because of their higher energy densities and better combustion efficiencies compared with ethanol. Microbial production of isopentenols has been developed recently via metabolically engineered E. coli. However, current yields remain low and the underlying pathways require systematic optimization.
  
  
  RESULTS
  In this study, we targeted the E. coli native 2-methyl-(D)-erythritol-4-phosphate (MEP) pathway and its upstream glycolysis pathway for the optimization of isopentenol production. Two codon optimized genes, nudF and yhfR from Bacillus subtilis, were synthesized and expressed in E. coli W3110 to confer the isopentenol production of the strain. Two key enzymes (IspG and Dxs) were then overexpressed to optimize the E. coli native MEP pathway, which led to a significant increase (3.3-fold) in isopentenol production. Subsequently, the glycolysis pathway was tuned to enhance the precursor and NADPH supplies for the MEP pathway by activating the pentose phosphate pathway (PPP) and Entner-Doudoroff pathway (ED), which resulted in additional 1.9 folds of increase in isopentenol production. A 5 L-scale batch cultivation experiment was finally implemented, showing a total of 61.9 mg L(-1) isopentenol production from 20 g L(-1) of glucose.
  
  
  CONCLUSIONS
  The isopentenol production was successfully increased through multi-step optimization of the MEP and its upstream glycolysis pathways. It demonstrated that the total fluxes and their balance of the precursors of the MEP pathway are of critical importance in isopentenol production. In the future, an elucidation of the contribution of PPP and ED to MEP is needed for further optimization of isopentenol production.
authors.forename: Huaiwei
authors.initials: H
authors.lastname: Liu
authors.forename: Yang
authors.initials: Y
authors.lastname: Wang
authors.forename: Qiang
authors.initials: Q
authors.lastname: Tang
authors.forename: Wentao
authors.initials: W
authors.lastname: Kong
authors.forename: Wook-Jin
authors.initials: WJ
authors.lastname: Chung
authors.forename: Ting
authors.initials: T
authors.lastname: Lu
mesh_terms: D061249:Batch Cell Culture Techniques; D053898:Biosynthetic Pathways; D003062:Codon / Q000235:genetics; D004896:Erythritol / Q000031:analogs & derivatives* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism; D005285:Fermentation; D006019:Glycolysis; D060847:Metabolic Engineering / Q000379:methods*; D000439:Pentanols / Q000378:metabolism*; D013403:Sugar Phosphates / Q000378:metabolism*; D013997:Time Factors
chemical_list: C114232:2-C-methylerythritol 4-phosphate; D003062:Codon; D029968:Escherichia coli Proteins; D000439:Pentanols; D013403:Sugar Phosphates; C056744:isopentenol; D004896:Erythritol