A yeast platform for high-level synthesis of tetrahydroisoquinoline alkaloids

Lauren Narcross,John E Dueber,Kaspar Kevvai,Brian Choi,Vincent J J Martin,Parbir S Grewal,Leanne Bourgeois,Michael E Pyne

doi:10.1038/s41467-020-17172-x

Abstract

The tetrahydroisoquinoline (THIQ) moiety is a privileged substructure of many bioactive natural products and semi-synthetic analogs. Plants manufacture more than 3,000 THIQ alkaloids, including the opioids morphine and codeine. While microbial species have been engineered to synthesize a few compounds from the benzylisoquinoline alkaloid (BIA) family of THIQs, low product titers impede industrial viability and limit access to the full chemical space. Here we report a yeast THIQ platform by increasing production of the central BIA intermediate (S)-reticuline to 4.6 g L−1, a 57,000-fold improvement over our first-generation strain. We show that gains in BIA output coincide with the formation of several substituted THIQs derived from amino acid catabolism. We use these insights to repurpose the Ehrlich pathway and synthesize an array of THIQ structures. This work provides a blueprint for building diverse alkaloid scaffolds and enables the targeted overproduction of thousands of THIQ products, including natural and semi-synthetic opioids.

Highlights

The tetrahydroisoquinoline (THIQ) moiety is a privileged substructure of many bioactive natural products and semi-synthetic analogs
To improve biosynthesis of THIQs in yeast, we focused on the first committed reaction and major rate-limiting step of the canonical benzylisoquinoline alkaloid (BIA) pathway
Our work demonstrates a major step towards industrial synthesis of microbially sourced THIQ pharmaceuticals by increasing production of the key BIA intermediate (S)-reticuline to 4.6 g L−1 using a simple mineral medium

Summary

Introduction

The tetrahydroisoquinoline (THIQ) moiety is a privileged substructure of many bioactive natural products and semi-synthetic analogs. Global demand for morphinan BIAs is presently met through extraction from opium poppy (Papaver somniferum), most BIAs do not accumulate to sufficient concentrations in plant tissues To begin exploring this untapped natural diversity, plant pathways mediating synthesis of noscapine, sanguinarine, morphine, codeine, and hydrocodone have been reconstructed in yeast[11,12,13,14]. Despite these achievements, present yeast BIA titers have been limited to

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Conclusion