(569f) Repurposing Chloramphenicol Acetyltransferase for a Robust Ester Biosynthesis Platform

Esters are versatile chemicals that have broad applications as flavors, fragrances, solvents, and potential drop-in biofuels. The key enzyme to synthesize esters is alcohol acyltransferase (AAT) that condenses alcohols and acyl-CoAs derived from cellular metabolism under a mild aqueous condition. Particularly, thermostable (≥ 50^oC) AAT plays a significant role for developing an industrial bioprocess to produce esters from lignocellulosic biomass but such an enzyme has not yet been reported to date. To enable a robust ester biosynthesis platform, we repurposed chloramphenicol acetyltransferase (CAT) to function like an AAT by harnessing its enzyme promiscuity. Through bioprospecting and rational protein design, we developed a highly thermostable CAT with a broad substrate range, which can function as an AAT for microbial whole cell biocatalyst development. The high thermostability and promiscuity of the engineered CAT make it an ideal platform for microbial biosynthesis of designer bioesters.