Production of medium chain length fatty alcohols from glucose in Escherichia coli

Abstract

Metabolic engineering offers the opportunity to produce a wide range of commodity chemicals that are currently derived from petroleum or other non-renewable resources. Microbial synthesis of fatty alcohols is an attractive process because it can control the distribution of chain lengths and utilize low cost fermentation substrates. Specifically, primary alcohols with chain lengths of 12 to 14 carbons have many uses in the production of detergents, surfactants, and personal care products. The current challenge is to produce these compounds at titers and yields that would make them economically competitive. Here, we demonstrate a metabolic engineering strategy for producing fatty alcohols from glucose. To produce a high level of 1-dodecanol and 1-tetradecanol, an acyl-ACP thioesterase (BTE), an acyl-CoA ligase (FadD), and an acyl-CoA/aldehyde reductase (MAACR) were overexpressed in an engineered strain of Escherichia coli. Yields were improved by balancing expression levels of each gene, using a fed-batch cultivation strategy, and adding a solvent to the culture for extracting the product from cells. Using these strategies, a titer of over 1.6 g/L fatty alcohol with a yield of over 0.13 g fatty alcohol / g carbon source was achieved. These are the highest reported yield of fatty alcohols produced from glucose in E. coli.

A Fatty Acyl Coenzyme A Reductase Promotes Wax Ester Accumulation in Rhodococcus jostii RHA1

“…Production, purification, and characterization of recombinant FcrA. We focused further efforts on exploring the function of FcrA, as related two-domain enzymes completely reduce acyl-CoAs to fatty alcohols (28) and have been used in biotechnology applications (31,32). To investigate the activity of FcrA, we produced it as a C-terminally His-tagged protein in RHA1 and purified it to greater than 95% apparent homogeneity (Fig.…”

Metabolic engineering of fatty acyl-ACP reductase-dependent pathway to improve fatty alcohol production in Escherichia coli

“…Interestingly, it was demonstrated that a fatty acyl-CoA reductase (MAACR) from Marinobacter aquaeolei VT8 reduces fatty acyl-CoAs into fatty alcohols in in vitro enzymatic assays, and fatty aldehydes were clearly intermediate products (Hofvander et al, 2011;Willis et al, 2011). The E. coli with overexpressed a modified thioesterase, FadD and MAACR (fatty acyl-CoA reductase) from M. aquaeolei VT8 produced 1.725 g/L fatty alcohols under the fermentation condition (Liu et al, 2013), and the other E. coli strain overexpressed an acyl-ACP thioesterase (BTE), FadD and the same fatty acyl-CoA reductase achieved 1.6 g/L fatty alcohols with yield of over 0.13 g fatty alcohol/g glucose (Youngquist et al, 2013). The fatty acyl-CoA reductase Acr1 from Acinetobacter calcoaceticus has been widely used for overproduction of fatty alcohols in E. coli.…”

Retro-biosynthetic screening of a modular pathway design achieves selective route for microbial synthesis of 4-methyl-pentanol

“…Although natural hosts and metabolic pathways have been used for decades in the production of chemical products, the desire to synthesize direct replacements for current petroleum products has led to appropriation of natural pathways for production of noncognate chemicals [1][2][3] . Early examples of this new paradigm have focused on introducing or combining portions of natural pathways in alternative host organisms or creating new products by altering the termination of natural pathways of a host organism with promiscuous enzymes 1,[3][4][5][6][7][8][9][10][11] . Recent work has been focused on improving modified natural pathways by substituting new enzymes to improve kinetics, improve expression or utilize alternate cofactors [12][13][14] .…”

“…Microbial synthesis of these reduced chemical species by de novo designed pathways can potentially lead to more efficient production strains, which are necessary for next-generation targets to achieve commercial relevance. Predominantly, these pathways have employed reconstitution of natural pathways in new hosts (acetone-butanol-ethanol 5,6 ) or modified termination of natural pathways (fatty acid synthesis (FAS) 11,23,24 , amino acid synthesis 12,28 , isoprenoid synthesis 22 ).…”