Heterologous Production of Isopropanol Using Metabolically Engineered Acetobacterium woodii Strains

Abstract

The depletion of fossil fuel resources and the CO2 emissions coupled with petroleum-based industrial processes present a relevant issue for the whole of society. An alternative to the fossil-based production of chemicals is microbial fermentation using acetogens. Acetogenic bacteria are able to metabolize CO or CO2 (+H2) via the Wood–Ljungdahl pathway. As isopropanol is widely used in a variety of industrial branches, it is advantageous to find a fossil-independent production process. In this study, Acetobacterium woodii was employed to produce isopropanol via plasmid-based expression of the enzymes thiolase A, CoA-transferase, acetoacetate decarboxylase and secondary alcohol dehydrogenase. An examination of the enzymes originating from different organisms led to a maximum isopropanol production of 5.64 ± 1.08 mM using CO2 + H2 as the carbon and energy source. To this end, the genes thlA (encoding thiolase A) and ctfA/ctfB (encoding CoA-transferase) of Clostridium scatologenes, adc (encoding acetoacetate decarboxylase) originating from C. acetobutylicum and sadH (encoding secondary alcohol dehydrogenase) of C. beijerinckii DSM 6423 were employed. Since bottlenecks in the isopropanol production pathway are known, optimization of the strain was investigated, resulting in a 2.5-fold increase in isopropanol concentration.

Engineered acetogenic bacteria as microbial cell factory for diversified biochemicals

“…The best-performing recombinant strain was able to produce 13.4 g/L of isopropanol in continuous gas fermentation. In addition, plasmid-based heterologous expression of the genes thlA-ctfAB-adc-sadh in A. woodii led to a maximum isopropanol production of 13.87 mM using CO 2 and H 2 as the carbon and energy source ( Höfele et al, 2023 ). Recent advancements by LanzaTech have demonstrated commercial-ready levels of acetone and isopropanol production using genomically integrated strains, with continuous production rates reaching up to ∼3 g/L/h ( Liew et al, 2022 ).…”