Engineering Escherichia coli for selective 1-decanol production using the reverse β-oxidation (rBOX) pathway

Journal Article · Fri Jul 21 00:00:00 EDT 2023 · Metabolic Engineering

DOI:https://doi.org/10.1016/j.ymben.2023.07.006· OSTI ID:2424752

Chen, Jing ^[1]; Gonzalez, Ramon ^[1]

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1. University of South Florida, St. Petersburg, FL (United States)

1-Decanol has great value in the pharmaceutical and fragrance industries and plays an important role in the chemical industry. Here, in this study, we engineered Escherichia coli to selectively synthesize 1-decanol by using enzymes of the core reverse β-oxidation (rBOX) pathway and termination module with overlapping chain-length specificity. Through screening for acyl-CoA reductase termination enzymes and proper regulation of rBOX pathway expression, a 1-decanol titer of 1.4 g/L was achieved. Further improvements were realized by engineering pyruvate dissimilation to ensure the generation of NADH through pyruvate dehydrogenase (PDH) and reducing byproduct synthesis via a tailored YigI thioesterase knockout, increasing 1-decanol titer to 1.9 g/L. The engineered strain produced about 4.4 g/L 1-decanol with a yield of 0.21 g/g in 36 hours in a bi-phasic fermentation that used a dodecane overlay to increase 1-decanol transport and reduce its toxicity. Adjustment of pathway expression (varying inducer concentration) and cell growth (oxygen availability) enabled 1-decanol production at 6.1 g/L (0.26 g/g yield) and 10.05 g/L (0.2 g/g yield) using rich in shake flasks and bioreactor, respectively. Remarkably, the use of minimal medium resulted in 1-decanol production at 100% specificity at 2.8 g/L (0.14 g/g yield) and a per cell mass yield higher than rich medium. These 1-decanol titers, yields and purity are at least 10-fold higher than others reported to date and the engineered strain shows great potential for industrial production. Taken together, our findings suggest that using rBOX pathway and termination enzymes of proper chain-length specificity in combination with optimal chassis engineering should be an effective approach for the selective production of alcohols.

• Research Organization: University of South Florida, St. Petersburg, FL (United States)
• Sponsoring Organization: National Science Foundation (NSF); USDOE
• OSTI ID: 2424752
• Alternate ID(s): OSTI ID: 2000220
• Journal Information: Metabolic Engineering, Journal Name: Metabolic Engineering Journal Issue: C Vol. 79; ISSN 1096-7176
• Publisher: Elsevier
• Country of Publication: United States
• Language: English

Related Subjects

1-decanol
59 BASIC BIOLOGICAL SCIENCES
Biotechnology & Applied Microbiology
reversal β-oxidation (rBOX)
selectively producing
termination enzyme