They’re a fungal fraternity known for their ‘milk’; Lactarius mushrooms exude colourful latex fluids when their flesh is damaged. In particular, the vibrant, orange-hued latex of Lactarius deliciosus—also known as the saffron milkcap mushroom—forms part of its defences. Intriguingly, this latex holds a variety of compounds called sesquiterpenes, which may not only help develop medicines and cosmetics, but may even be used as jet fuel.
To enable more industrial applications for sesquiterpenes, it’s critical to get a clearer picture of the molecular machines that help produce them in nature: enzymes called sesquiterpene synthases (STSs), said Yee Hwee Lim, Director of the Chemical Biotechnology and Biocatalysis division at A*STAR’s Institute of Sustainability for Chemicals, Energy and Environment (ISCE2).
“Currently, scientists still can’t accurately predict the products of STSs,” Lim explained, adding that attempts to unravel the enzymes’ functions with conventional methods have been slow and costly.
However, innovative genomic and bioinformatic approaches could shed more light on the mechanics of how STSs form sesquiterpenes. This would enable researchers to both predict thousands of naturally-occurring, yet undiscovered STS products; and to engineer STSs that efficiently and sustainably produce sesquiterpenoids of practical use, said Lim.
In partnership with Congqiang Zhang, a Principal Investigator at A*STAR’s Singapore Institute of Food and Biotechnology Innovation (SIFBI), and other collaborators from ISCE2, SIFBI and the University of Illinois, US, Lim’s team embarked on comprehensive analyses to characterise STSs made by L. deliciosus.
First, the researchers hunted for clues in the mushroom’s genome, using bioinformatics to predict the likeliest genes behind STS production. To validate their predictions, these genes were then expressed in a specially-engineered strain of Escherichia coli bacteria.
“With our proprietary E. coli platform, which is efficient and easily scalable, we can essentially produce any terpene from fungi and plants,” said Zhang.
The STSs they made were then analysed using gas chromatography-mass spectrometry to determine their chemical compositions. Excitingly, this led to the discovery of the first member of a previously-unknown STS clade which creates unique terpene ‘scaffolds’: spiralling loops held together by three interlocked rings.
A phylogenetic analysis—akin to a family tree—also offered clues on the evolutionary relationships between the new clade and other known STSs. “Although we’ve been working on fungal terpenoids for about a decade, this study revealed that there are still many unknowns in their biosynthesis,” Lim said.
The research group continues to explore uncharted territory in novel STSs and their potential applications. At present, they have filed two patents for the use of their newly discovered STS in cosmetic fragrances and biofuels, leveraging the compound’s energy-rich hydrocarbon properties.
The A*STAR-affiliated researchers contributing to this research are from the Institute of Sustainability for Chemicals, Energy and Environment (ISCE2) and the Singapore Institute of Food and Biotechnology Innovation (SIFBI).
