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Sunstrum, Frederick Guy

University of British Columbia

The plant kingdom has always been a useful source of therapeutic molecules for humans. Montbretin A (MbA) is a complex acylated flavonol glycoside found in the underground storage organs, called corms, of the ornamental plant montbretia (Crocosmia x crocosmiiflora). MbA was discovered as a highly efficient and specific inhibitor of the human pancreatic α-amylase (HPA), making it an attractive candidate for the treatment of type 2 diabetes, a worldwide epidemic. The original plant source montbretia is not adapted for large scale cultivation, and MbA’s complex structure renders chemical synthesis infeasible for scalable production. Therefore, we are aiming to engineer a heterologous system for large-scale sustainable MbA production for treatment of type 2 diabetes. To this end we are using synthetic biology to develop an MbA producing Nicotiana benthamiana (tobacco) strain. This is possible due to the recent complete elucidation of the MbA biosynthetic pathway, including all essential genes and enzymes from montbretia. Small amounts of MbA can be produced in N. benthamiana through transient expression of the MbA biosynthetic enzymes. However, much higher quantities of montbretin B (MbB) and montbretin C (MbC), which differ from MbA in the identity of their acyl groups, were produced. MbB and MbC possess a coumaroyl and a feruloyl group, respectively, and are not effective HPA inhibitors. To influence acyl group availability, specifically increase the amount of caffeoyl-CoA for MbA biosynthesis, we investigated a set of enzymes involved in caffeoyl-CoA biosynthesis in montbretia, specifically 4-coumaroyl CoA ligases (4CL), hydroxycinnamoyl- CoA shikimate/quinate hydroxycinnamoyltransferases (HCT), 4-coumarate 3-hydroxylases (C3Hs), and caffeoyl-shikimate esterases (CSE). Co-expression of those caffeoyl-CoA biosynthetic genes together with MbA biosynthetic genes resulted in up to 10-fold higher MbA yields.

Text

2021-12-07

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/80382

Genome Science and Technology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/