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Elucidation of the biosynthesis pathway of the glaucoma drug pilocarpine in Pilocarpus plants


Pilocarpine is a plant-derived alkaloid used to treat primary angle closure glaucoma and dry mouth. To meet the commercial demand for pilocarpine, the alkaloid is extracted from wild Pilocarpus plants in the Amazon rainforest. Excessive harvesting of wild populations has led to the official listing of several Pilocarpus species as threatened. Synthetic biology offers a sustainable approach for pilocarpine production. However, to produce the compound in heterologous hosts such as yeast, the biosynthesis pathway of pilocarpine needs to be elucidated first. In PiloSyn, I aim to unravel the natural biosynthesis pathway of pilocarpine, which is currently completely unknown. My host group recently obtained transcriptomics and metabolomics data sets of two Pilocarpus species, and these will serve as the basis of the project. As several pathway options are conceivable for pilocarpine biosynthesis, I will first reduce the number of options by identifying the metabolic precursors of the pathway and by obtaining structural and spatial information about possible pathway intermediates. Then, using the transcriptomics data sets, I will identify the enzymes that catalyze each of the pathway reactions. For this, I will use a learning-by-constructing approach in transiently transformed leaves of Nicotiana benthamiana. The results of PiloSyn can be used to establish a cheap and sustainable pilocarpine source in a heterologous host. This will eliminate the need to harvest wild Pilocarpus plants, thus is preventing their extinction and protecting the fragile ecosystem of the Amazon rainforest. Through PiloSyn, I will expand my knowledge on enzyme discovery and pathway elucidation and receive valuable training to become an independent and mature researcher in plant metabolism.

Funding Scheme



Net EU contribution
€ 214 934,40
Norregade 10
1165 Kobenhavn

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Danmark Hovedstaden Byen København
Activity type
Higher or Secondary Education Establishments
EU contribution
No data