β-Amyrin synthase from Conyza blinii expressed in Saccharomyces cerevisiae

Conyza blinii H.Lev. is a widely used medicinal herb in southwestern China. The main pharmacological components of C. blinii are a class of oleanane-type pentacyclic triterpene glycosides known as conyzasaponins, which are thought to be synthesized from β-amyrin. However, no genes involved in the conyzasaponin pathway have previously been identified. Here, we identify an oxidosqualene cyclase (OSC), a β-amyrin synthase, which mediates cyclization of 2,3-oxidosqualene to yield β-amyrin. Ten OSC sequences were isolated from C. blinii transcript tags. Phylogenetic analysis was used to select the tag Cb18076 as the putative β-amyrin synthase, named CbβAS. The open reading frame of CbβAS is 2286 bp and encodes 761 amino acids. Its mature protein contains the highly conserved motifs (QXXXGXW/DCTAE) of OSCs and (MWCYCR) of β-amyrin synthases. Transcription of CbβAS was upregulated 4-24 h after treatment of the seedlings of the C. blinii cultivar with methyl jasmonate. Furthermore, expression of CbβAS in Saccharomyces cerevisiae successfully yielded β-amyrin. The chemical structures and concentrations of β-amyrin were confirmed by GC-MS/MS. The target yeast ultimately produced 4.432 mg·L-1 β-amyrin. Thus, CbβAS is an OSC involved in conyzasaponin biosynthesis.