The 4-year, EU-funded TriForC (Triterpene’s For Commercialisation) project has come to a close, resulting in the publication of the most extensive and significant algal strain engineering study to date
Algenuity, the specialist algal biology company based in Stewartby, UK, was one of 11 international partners, representing eight countries, in the consortium. The aim of the project was to develop synthetic biology methods to make plant triterpenes in a sustainable way, using algae and yeast as host organisms.
Triterpenes belong to a diverse class of organic C30 compounds that generally comprise five connected rings, elaborated by modifications such as methylation, oxidation and glycosylation.
They play a major role in plant defence mechanisms and frequently form the basis of herbal medicines with some promise shown as potential pharmaceuticals because of their potent bioactivities.
However, they are produced in low levels by their natural hosts and it is often difficult to identify and then isolate the bioactive component from the complicated mixture of compounds within a plant.
The aim of the TriForC project was to use a synthetic biology platform to find an environmentally viable way of producing these compounds in industrially relevant microbial systems for use in biopharmaceuticals (and, perhaps, biopesticides).1
Andrew Spicer, CEO of Algenuity, explains: “The concept of the project was to use omics-based approaches to identify the enzymes within biochemical pathways and then synthetic biology methods to express those enzymes in both yeast and algae chassis organisms."
"Algae were promising because they are more closely related to plants than yeast, so we could predict that they might be more tolerant of the engineering modifications we were making. We looked at several different algal strains and the most successful was a diatom called Phaeodactylum tricornutum, which has already been used as a microalgal chassis in synthetic biology.”
“Sophisticated tools to control expression in this strain already existed — including promoters, terminators and control elements — and the strain’s phylogeny means that it tends to be easier to successfully express heterologous genes."
"We introduced a biosynthetic pathway into P. tricornutum to produce a specific triterpene called betulin, which has previously shown potential as a pharmaceutical precursor for the treatment of certain cancers and HIV,” he added.
The Algenuity team was able to express five genes in parallel — two selectable markers and three genes — which is a significant outcome considering that most groups aim to express only one gene for a given project.
An oxidosqualene cyclase scaffold enzyme was expressed to convert a native substrate into a triterpene scaffold, whereas cytochrome P450 mono-oxygenase and reductases were expressed to promote desired modifications to the scaffold.
Andrew concluded: “We conducted a great deal of research to find and code the equivalent genes for enzymes from different plants to give us the best combination of productivity and specificity for our target compound.”
“The success of this project clearly demonstrates that metabolic engineering is possible in algae. We can now look at applying this technology to other strains and grow them to much higher densities, which could be very promising for commercial scale-up. We are excited to see where this leads us and others in future studies.”
1. S. D’Adamo, et al., “Engineering the Unicellular Alga Phaeodactylum tricornutum for High-Value Plant Triterpenoid Production,” Plant Biotechnology Journal (2018): doi: 10.1111/pbi.12948.