Unravelling the hidden power of esterases for biomanufacturing of short-chain esters.

Microbial production of esters has recently garnered wide attention, but the current production metrics are low. Evidently, the ester precursors (organic acids and alcohols) can be accumulated at higher titers by microbes like Escherichia coli. Hence, we hypothesized that their 'direct esterification' using esterases will be efficient. We engineered esterases from various microorganisms into E. coli, along with overexpression of ethanol and lactate pathway genes. High cell density fermentation exhibited the strains possessing esterase-A (SSL76) and carbohydrate esterase (SSL74) as the potent candidates. Fed-batch fermentation at pH 7 resulted in 80 mg/L of ethyl acetate and 10 mg/L of ethyl lactate accumulation by SSL76. At pH 6, the total ester titer improved by 2.5-fold, with SSL76 producing 225 mg/L of ethyl acetate, and 18.2 mg/L of ethyl lactate, the highest reported titer in E. coli. To our knowledge, this is the first successful demonstration of short-chain ester production by engineering 'esterases' in E. coli.