Expanding ester biosynthesis in Escherichia coli

Abstract

To expand the capabilities of whole-cell biocatalysis, we have engineered Escherichia coli to produce various esters. The alcohol O-acyltransferase (ATF) class of enzyme uses acyl-CoA units for ester formation. The release of free CoA upon esterification with an alcohol provides the free energy to facilitate ester formation. The diversity of CoA molecules found in nature in combination with various alcohol biosynthetic pathways allows for the biosynthesis of a multitude of esters. Small to medium volatile esters have extensive applications in the flavor, fragrance, cosmetic, solvent, paint and coating industries. The present work enables the production of these compounds by designing several ester pathways in E. coli. The engineered pathways generated acetate esters of ethyl, propyl, isobutyl, 2-methyl-1-butyl, 3-methyl-1-butyl and 2-phenylethyl alcohols. In particular, we achieved high-level production of isobutyl acetate from glucose (17.2 g l−1). This strategy was expanded to realize pathways for tetradecyl acetate and several isobutyrate esters.

Identification and kinetics characterization of a wax ester hydrolase from a feather-degrading actinomycete

“…http://dx.doi.org/10.1101/178673 doi: bioRxiv preprint first posted online Aug. 21, 2017; biosynthesis of wax esters (Heilmann et al 2012;Rodriguez et al 2014;Santala et al 2014;Trani et al 1991;Tsujita et al 1999), but studies focused on the hydrolysis of wax esters have been few. The literature contains a few examples of wax ester hydrolases (EC 3.1.1.50) from plant (Huang et al 1978;Kalinowska and Wojciechowski 1985;Moreau and Huang 1981), fungal (Brahimihorn et al 1989), marine Kayama et al 1979;Mankura et al 1984;Patton et al 1975), and bacterial (Kalscheuer and Steinbuchel 2003) origins have previously been reported, however kinetic information is scant (Huang et al 1978;Kalinowska and Wojciechowski 1985).…”

Production of medium-chain, a, omega-bifunctional monomers from fatty acids and n-alkanes

“…In vivo esterification of fatty acids results in products with a low solubility, which can facilitate product removal and product inhibition 84,85,121 . Highest titer of ethyl ester was reached when nonanoic acid was applied as substrate, resulting in 0.82 mM of ethyl nonanoate from 1 mM nonanoic acid.…”

“…This strain expresses Eeb1 as alcohol acyltransferase. MDE3 did not produce alkyl alkanoates (Figure 5.6f), since Eeb1 is specific for short alcohols 85,93 . Ethyl ester synthesis was more efficient, and as a consequence more mono-ethyl dicarboxylic acid accumulated, up to 0.87 mM from noctane.…”

“…Of these AATs, AtfA and Eeb1 have a substrate specificity that enables short-chain alcohols to be coupled with medium-chain acyl-CoAs. Both enzymes have already been applied for (medium-chain) ester production in E. coli [83][84][85]93 . Hence, converting the MCFA to an ester should be possible.…”

“…In vivo ester production has gained a lot of interest, with the intention to produce biodiesel or fuels from fatty acids or carbohydrates [83][84][85]88,89,135,136 . AtfA and Eeb1 93 were applied to produce a variety of esters, including ethyl esters.…”