Engineering biosynthesis of the anticancer alkaloid noscapine in yeast

Abstract

Noscapine is a potential anticancer drug isolated from the opium poppy Papaver somniferum, and genes encoding enzymes responsible for the synthesis of noscapine have been recently discovered to be clustered on the genome of P. somniferum. Here, we reconstitute the noscapine gene cluster in Saccharomyces cerevisiae to achieve the microbial production of noscapine and related pathway intermediates, complementing and extending previous in planta and in vitro investigations. Our work provides structural validation of the secoberberine intermediates and the description of the narcotoline-4′-O-methyltransferase, suggesting this activity is catalysed by a unique heterodimer. We also reconstitute a 14-step biosynthetic pathway of noscapine from the simple alkaloid norlaudanosoline by engineering a yeast strain expressing 16 heterologous plant enzymes, achieving reconstitution of a complex plant pathway in a microbial host. Other engineered yeasts produce previously inaccessible pathway intermediates and a novel derivative, thereby advancing protoberberine and noscapine related drug discovery.

Engineering the biocatalytic selectivity of iridoid production in Saccharomyces cerevisiae

“…Double two e − oxidation catalyzed by geraniol oxidoreductase (GOR) afford the dialdehyde 8-oxogeranial 2 , which then undergoes reductive cyclization by iridoid synthase (ISY) to yield the bicyclic nepetalactol 3 (Geu-Flores et al, 2012). Production of the iridoid scaffold appears concise compared to other plant pathways that have recently been reconstituted in yeast, such as those producing sesquiterpenoids (Paddon et al, 2013) and benzylisoquinoline alkaloids (Galanie et al, 2015; Li and Smolke, 2016). However, building iridoid (and in turn MIA) “platform strains” (Nielsen, 2015) has proven to be challenging with each of the four enzymatic steps posing its own metabolic engineering obstacles.…”

Metabolic Engineering of Microorganisms for the Production of Natural Compounds

“…For example, phthalideisoquinoline noscapine pathway has been recently fully identified, along with the very recent success in noscapine biosynthesis in yeast (Figure 17). [276] The corresponding biosynthetic pathway also stems from S-scoulerine, sharing the common precursor with www.adv-biosys.com www.advancedsciencenews.com protoberberine alkaloids. However, microbial expression of the heterologous plant enzymes still retain a number of challenges endowed by spontaneous side reactions due to enzyme promiscuity, low efficiency, enzyme spatial localization, etc.…”

Synthetic Biology—The Synthesis of Biology

“…[117] The transfer of these biosynthetic pathways into industrially relevant producer cell hosts requires further adaptation and optimization steps.T his effort resulted in the successful production of therapeutic drugs in host cells,s uch as anticancer compounds (e.g.t axadiene [118] and noscapine), [119] drugs (e.g.artemisinin), [120] and narcotics (opioids). [117] The transfer of these biosynthetic pathways into industrially relevant producer cell hosts requires further adaptation and optimization steps.T his effort resulted in the successful production of therapeutic drugs in host cells,s uch as anticancer compounds (e.g.t axadiene [118] and noscapine), [119] drugs (e.g.artemisinin), [120] and narcotics (opioids).…”