Molecular Cloning and Bacterial Expression of Germacrene A Synthase cDNA from Crepidiastrum sonchifolium

Germacrene A synthase(GAS) catalyzes the biosynthesis of germacrene A, which is a key precursor for sesquiterpene lactones. Cloning of a novel full-length cDNA encoding GAS from the medicinal plant Crepidiastrum sonchifolium (designated CsGAS) is reported in this study. The cDNA is 1837 bp long and contains a 1680-bp open reading frame encoding a 559 amino-acid protein. The functional expression of the cDNA in Escherichia coli, as an N-terminal thioredoxin fusion protein, with the pET32a vector yielding a recombinant enzyme. Sequence analysis was used to compare this enzyme with the mechanistically related epi-aristolochene synthase from tobacco, and the effect of possible involvement of a number of amino acids in sesquiterpene synthase on product specificity was also discussed.