Metabolic Engineering of Fatty Acyl-ACP Reductase-Dependent Pathway to Improve Fatty Alcohol Production in Escherichia coli

Fatty alcohols are important components of surfactants and cosmetic products. The production of fatty alcohols from sustainable resources using microbial fermentation could reduce dependence on fossil fuels and greenhouse gas emission. However, the industrialization of this process has been hampered by the current low yield and productivity of this synthetic pathway.
As a result of metabolic engineering strategies, an Escherichia coli mutant containing Synechococcus elongatus fatty acyl-ACP reductase showed improved yield and productivity. This pathway avoids the extra ATP consuming either for synthesis of fatty acyl-CoAs or for activation of the fatty acids.
Proteomics analysis and in vitro enzymatic assays showed that endogenous E. coli AdhP is a major contributor to the reduction of fatty aldehydes to fatty alcohols.When the yield of fatty acyl-ACPs is sufficient as a result of engineering of AAR, the next useful modification to attempt would be improvement of AdhP activity. This also suggests that, if we want to overproduce fatty alkanes, we should knock out adhP and other homologous genes in E. coli.
Both in vitro and in vivo results clearly demonstrated that the activity and expression level of fatty acyl-CoA/ACP reductase is the rate-limiting step in the current protocol, increasing the fatty acyl-CoA/ACP reductase expression level, improving the activity of fatty acyl-CoA/ACP reductase, or identifying a better fatty acyl-ACP reductase to replace Synechococcus elongatus fatty acyl-ACP reductase in the future are promising avenues to realize industrialization of fatty alcohol biosynthesis.
In 2.5-L fed-batch fermentation with glycerol as the only carbon source, the most productive E. coli mutant produced
0.75 g/L fatty alcohols (0.02 g fatty alcohol/g glycerol) with a productivity of up to 0.06 g/L/h.
Our findings highlight the value of rational metabolic engineer-ing of the complete fatty alcohol biosynthetic pathway and provide direction for improving the yield of fatty alcohols. Further investigation is warranted to understand and engineer the regulatory interactions between biosynthetic intermediates, final products, and individual enzymes in the E. coli pathway. More generally, further efforts on both fronts are warranted, and our results will accelerate the pace of realizing industrial production of fatty alcohols.