Functional Genomics and Metabolite Profiling as Tools for Alkaloid Biosynthetic Gene Discovery

Abstract

The benzylisoquinoline alkaloids (BIAs) are diverse group of plant specialized metabolites found in the families Papaveracea, Ranunculaceae, Berberidaceae and Menispermaceae. Plants remain the only commercial source for BIAs and their biosynthesis is poorly understood. O-methyltransferases (OMTs) are wide spread in BIA biosynthesis. Putative OMTs were found in stem and root Next-Generation Sequencing transcriptomic databases. Putative OMT cDNAs were isolated from Papaver somniferum and commercially synthesized. Recombinant protoberberine 2-O-methyltransferase (2OMT) was heterologously expressed in Escherichia coli and assayed. 2OMT demonstrated the 2-O-methylation of protoberberine alkaloids and the 7-O-methylation of simple BIAs. The substrate range and tissue specific expression of 2OMT suggest its in vivo role is converting (S)-cheilanthifoline to (S)-sinactine. A LC-MS based targeted alkaloid profiling of twenty BIA producing species from the families Papaveracea, Ranunculaceae, Berberidaceae and Menispermaceae was conducted.