Production of the Inaccessible Sesquiterpene (−)‐5‐Epieremophilene by Metabolically Engineered Escherichia coli

(?)‐5‐Epieremophilene, an epimer of the versatile sesquiterpene (+)‐valencene, is an inaccessible natural product catalyzed by three sesquiterpene synthases (SmSTPSs1‐3) of the Chinese medicinal herb Salvia miltiorrhiza, and its biological activity remains less explored. In this study, three metabolically engineered Escherichia coli strains were constructed for (?)‐5‐epieremophilene production with yields of 42.4–76.0 mg/L in shake‐flask culture. Introducing an additional copy of farnesyl diphosphate synthase (FDPS) gene through fusion expression of SmSTPS1‐FDPS or dividing the FDP synthetic pathway into two modules resulted in significantly improved production, and ultimately 250 mg of (?)‐5‐epieremophilene were achieved. Biological assay indicated that (?)‐5‐epieremophilene showed significant antifeedant activity against Helicoverpa armigera (EC₅₀=1.25 μg/cm²), a common pest of S. miltiorrhiza, implying its potential defensive role in the plant. The results provided an ideal material supply for studying other potential biological activities of (?)‐5‐epieremophilene, and also a strategy for manipulating terpene production in engineered E. coli using synthetic biology.