Trichloroacetic acid-facilitated coenzyme A-SH-stabilization-based end-point spectrophotometric 5,5 -dithiobis-2-nitrobenzoic acid assay for plant terpene alcohol acetyltransferases

Abstract

Acyltransferases catalyze critical acylation of metabolic intermediates in metabolic pathways and are important in metabolome diversiWcation in secondary metabolism. Terpenes form the largest and most diverse Wed group of secondary metabolites (>35,000 compounds). Terpene alcohol acetyltransferases (AATs)1 are responsible for biogeneration of volatile esters that attribute unique and Wne olfactory aromas to herbs, Xowers, fruits, and foods. AATs catalyze transfer of an acyl group from acetyl/acyl coenzyme A to an alcohol, stoichiometrically generating ester and coenzyme A-SH (CoASH). Recently, the signiWcance of AATs in the Xavor and fragrance of phytogenic foods, drinks, and aromatic oils and in developmental physiology of Xowers and fruits has attracted their biochemical characterization in plants [1–5] for metabolic engineering initiatives. Radioactivity [5] or GC-based quantitation [1,3,5] of ester formed by the reaction are employed to monitor the enzyme activity.