Engineering a microbial biosynthesis platform for de novo production of tropane alkaloids

Prashanth Srinivasan,Christina D. Smolke

doi:10.1038/s41467-019-11588-w

Prashanth Srinivasan, Christina D. Smolke

Open Access

https://doi.org/10.1038/s41467-019-11588-w

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Abstract

Tropane alkaloids (TAs) are a class of phytochemicals produced by plants of the nightshade family used for treating diverse neurological disorders. Here, we demonstrate de novo production of tropine, a key intermediate in the biosynthetic pathway of medicinal TAs such as scopolamine, from simple carbon and nitrogen sources in yeast (Saccharomyces cerevisiae). Our engineered strain incorporates 15 additional genes, including 11 derived from diverse plants and bacteria, and 7 disruptions to yeast regulatory or biosynthetic proteins to produce tropine at titers of 6 mg/L. We also demonstrate the utility of our engineered yeast platform for the discovery of TA derivatives by combining biosynthetic modules from distant plant lineages to achieve de novo production of cinnamoyltropine, a non-canonical TA. Our engineered strain constitutes a starting point for future optimization efforts towards realizing industrial fermentation of medicinal TAs and a platform for the synthesis of TA derivatives with enhanced bioactivities.

Highlights

Tropane alkaloids (TAs) are a class of phytochemicals produced by plants of the nightshade family used for treating diverse neurological disorders
The tropine moiety of TAs is derived from arginine via putrescine, a polyamine required for ribosome biogenesis and mRNA translation[24]
We focused on engineering a putrescine-overproducing strain by overexpressing native genes involved in arginine metabolism and polyamine biosynthesis (Fig. 2a)

Summary

Introduction

Tropane alkaloids (TAs) are a class of phytochemicals produced by plants of the nightshade family used for treating diverse neurological disorders. We demonstrate de novo production of tropine, a key intermediate in the biosynthetic pathway of medicinal TAs such as scopolamine, from simple carbon and nitrogen sources in yeast (Saccharomyces cerevisiae). Our engineered strain incorporates 15 additional genes, including 11 derived from diverse plants and bacteria, and 7 disruptions to yeast regulatory or biosynthetic proteins to produce tropine at titers of 6 mg/L. Researchers reported the discovery of an unusual type III polyketide synthase (PKS) from Atropa belladonna, which condenses two malonyl-CoAs with NMPy in the absence of an additional starter unit to form 4-(1-methyl-2-pyrrolidinyl)-3-oxobutanoic acid (MPOB), and a cytochrome P450, which catalyzes the oxidation and cyclization of MPOB to tropinone[19], thereby completing the biosynthetic pathway between arginine and tropine (Fig. 1)

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