Microbial Cell Factories for Plant Secondary Metabolites - Engineering Glucosinolates in Bacteria and Yeast

Glucosinolates are amino acid-derived sulphur-rich specialised metabolites characteristic of the Brassicales order and well known for their health-promoting effects upon intake of e.g. broccoli and cabbage. However, high amounts of broccoli and cabbage must be eaten on a regular basis to fully realize the health-beneficial effects and the current supplement market is based on products with unreliable amounts, if any at all. This has primed a strong desire to develop an efficient microbial cell factory for glucosinolate production as stable, rich source.

Building upon our long lasting experience with glucosinolate pathway reconstruction and enzyme discovery, we have moved onto engineering the synthetic pathway in Escherichia coli and Saccharomyces cerevisiae. We have successfully established the production of benzyl glucosinolate (BGLS)¹ and indol-3-ylmethyl (I3M)² glucosinolate as well as we have made substantial advancements in the amino acid chain elongation^3,4, which is a necessary first step for some of the most potent glucosinolates. We were able to boost the synthesis of BGLS, by systematic pathway engineering, optimization of the flux through the entry point cytochrome P450 enzymes and the last sulfotransferase step.

Towards our goal of engineering microbial high-level production of glucosinolates, we have encountered challenges related to the microbial sulphur chemistry to produce the sulphur rich glucosinolates and to premature abortion of the pathway by the intermediates escaping from the host cells. We apply pathway engineering as well as transport engineering approaches to overcome these challenges. Learnt lessons will be discussed.

1. Petersen, Metabolic engineering (2019)
2. Mikkelsen, Metabolic engineering (2012)
3. Mirza, Metabolic engineering (2016)
4. Petersen, Bioscience reports (2019)