Functional Expression and Characterization of Trichome-Specific (-)-Limonene Synthase and (+)-α-Pinene Synthase fromCannabis sativa

Abstract

Two recombinant, stereospecific monoterpene synthases, a (-)-limonene synthase (CsTPS1) and a (+)-α-pinene synthase (CsTPS2), encoded by Cannabis sativa L. cv. ‘Skunk’ trichome mRNA, have been isolated and characterized. Recombinant CsTPS1 showed a Kmvalue of 6.8 μM, a Vmaxof 1.1 × 10−4μmol/min and Vmax/Kmof 0.016; the pH optimum was determined at pH 6.5, and a temperature optimum at 40°C. Recombinant CsTPS2 showed a Kmvalue of 10.5 μM, a Vmaxof 2.2 × 10−4μmol/min and Vmax/Kmof 0.021; the pH optimum was determined at pH 7.0, and a temperature optimum at 30°C. Phylogenetic analysis showed that both CsTPSs group within the angiosperms and belong to the Tpsb subgroup of monoterpene synthases. The enzymatic products (-)-limonene and (+)-α-pinene were detected as natural products in C. sativa trichomes.