title: Rapid ester biosynthesis screening reveals a high activity alcohol-O-acyltransferase (AATase) from tomato fruit.
abstract: Ethyl and acetate esters are naturally produced in various yeasts, plants, and bacteria. The biosynthetic pathways that produce these esters share a common reaction step, the condensation of acetyl/acyl-CoA with an alcohol by alcohol-O-acetyl/acyltransferase (AATase). Recent metabolic engineering efforts exploit AATase activity to produce fatty acid ethyl esters as potential diesel fuel replacements as well as short- and medium-chain volatile esters as fragrance and flavor compounds. These efforts have been limited by the lack of a rapid screen to quantify ester biosynthesis. Enzyme engineering efforts have also been limited by the lack of a high throughput screen for AATase activity. Here, we developed a high throughput assay for AATase activity and used this assay to discover a high activity AATase from tomato fruit, Solanum lycopersicum (Atf-S.l). Atf1-S.l exhibited broad specificity towards acyl-CoAs with chain length from C4 to C10 and was specific towards 1-pentanol. The AATase screen also revealed new acyl-CoA substrate specificities for Atf1, Atf2, Eht1, and Eeb1 from Saccharomyces cerevisiae, and Atf-C.m from melon fruit, Cucumis melo, thus increasing the pool of characterized AATases that can be used in ester biosynthesis of ester-based fragrance and flavor compounds as well as fatty acid ethyl ester biofuels.
authors.affiliation: Chemical and Environmental Engineering, University of California, Riverside, CA, USA.
authors.forename: Jyun-Liang
authors.initials: JL
authors.lastname: Lin
authors.affiliation: Biochemistry, University of California, Riverside, CA, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhu
authors.affiliation: Chemical and Environmental Engineering, University of California, Riverside, CA, USA. iwheeldon@engr.ucr.edu.
authors.forename: Ian
authors.initials: I
authors.lastname: Wheeldon
mesh_terms: D000105:Acetyl Coenzyme A / Q000737:chemistry; D000214:Acyl Coenzyme A / Q000737:chemistry; D004952:Esters / Q000378:metabolism; D005227:Fatty Acids / Q000096:biosynthesis*; D057166:High-Throughput Screening Assays / Q000379:methods*; D018551:Solanum lycopersicum / Q000201:enzymology*; D060847:Metabolic Engineering / Q000379:methods; D010940:Plant Proteins / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D000214:Acyl Coenzyme A; D004952:Esters; D005227:Fatty Acids; D010940:Plant Proteins; D011506:Proteins; D000105:Acetyl Coenzyme A; C079238:alcohol O-acetyltransferase
keywords: Acetylation; Alcohol-O-acetyl/acyltransferase (AATase); Ester biosynthesis; High throughput screening