title: Engineering an Escherichia coli platform to synthesize designer biodiesels.
abstract: Biodiesels, fatty acid esters (FAEs), can be synthesized by condensation of fatty acid acyl CoAs and alcohols via a wax ester synthase in living cells. Biodiesels have advantageous characteristics over petrodiesels such as biodegradability, a higher flash point, and less emission. Controlling fatty acid and alcohol moieties are critical to produce designer biodiesels with desirable physiochemical properties (e.g., high cetane number, low kinematic viscosity, high oxidative stability, and low cloud point). Here, we developed a flexible framework to engineer Escherichia coli cell factories to synthesize designer biodiesels directly from fermentable sugars. In this framework, we designed each FAE pathway as a biodiesel exchangeable production module consisting of acyl CoA, alcohol, and wax ester synthase submodules. By inserting the FAE modules in an engineered E. coli modular chassis cell, we generated E. coli cell factories to produce targeted biodiesels (e.g., fatty acid ethyl (FAEE) and isobutyl (FAIbE) esters) with tunable and controllable short-chain alcohol moieties. The engineered E. coli chassis carrying the FAIbE production module produced 54mg/L FAIbEs with high specificity, accounting for>90% of the total synthesized FAEs and ∼4.7 fold increase in FAIbE production compared to the wildtype. Fed-batch cultures further improved FAIbE production up to 165mg/L. By mixing ethanol and isobutanol submodules, we demonstrated controllable production of mixed FAEEs and FAIbEs. We envision the developed framework offers a flexible, alternative route to engineer designer biodiesels with tunable and controllable properties using biomass-derived fermentable sugars.
authors.affiliation: Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, Tennessee, United States.
authors.forename: Michael
authors.initials: M
authors.lastname: Wierzbicki
authors.affiliation: Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, Tennessee, United States.
authors.forename: Narayan
authors.initials: N
authors.lastname: Niraula
authors.affiliation: Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, Tennessee, United States.
authors.forename: Akshitha
authors.initials: A
authors.lastname: Yarrabothula
authors.affiliation: Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, Tennessee, United States; Bioenergy Science Center (BESC), Oak Ridge National Laboratory, Oak Ridge, Tennessee, United States.
authors.forename: Donovan S
authors.initials: DS
authors.lastname: Layton
authors.affiliation: Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, Tennessee, United States; Bredesen Center for Interdisciplinary Research and Graduate Education, The University of Tennessee, Knoxville, Tennessee, United States; Bioenergy Science Center (BESC), Oak Ridge National Laboratory, Oak Ridge, Tennessee, United States. Electronic address: ctrinh@utk.edu.
authors.forename: Cong T
authors.initials: CT
authors.lastname: Trinh
mesh_terms: D061249:Batch Cell Culture Techniques; D056804:Biofuels / Q000382:microbiology*; D018533:Biomass; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development*; D005227:Fatty Acids / Q000737:chemistry; D005233:Fatty Alcohols / Q000378:metabolism; D005285:Fermentation; D060847:Metabolic Engineering / Q000379:methods*
chemical_list: D056804:Biofuels; D005227:Fatty Acids; D005233:Fatty Alcohols
keywords: Advanced biofuels; Biodiesel modules; Designer biodiesels; Fatty acid ethyl esters; Fatty acid isobutyl esters; Modular cell chassis