Date of Defense
4-16-2024
Date of Graduation
4-2024
Department
Biological Sciences
First Advisor
Todd Barkman
Second Advisor
Frederick Stull
Abstract
Caffeine has evolved independently at least six times in at least five angiosperm lineages. Among these lineages, different species utilize unique combinations of genes and biosynthetic pathways to produce caffeine. Ilex paraguariensis (yerba mate) is a caffeine producing species of plant from the holly genus, Ilex and the family, Aquifoliaceae. It is native to South America where it is widely known for its use in a caffeine rich beverage called maté. Surprisingly, caffeine production is a recently evolved trait in yerba mate and little is known about how its complete caffeine biosynthetic pathway formed. The purpose of the present study was to take a closer look at the unique mechanisms that underlie caffeine production in yerba mate through the study of its ancestral caffeine synthase enzymes. To do so, we performed ancestral sequence reconstruction to predict the sequences of two ancient genes encoding caffeine synthase enzymes in yerba mate. The function of these enzymes were then characterized via radioactive assays and high performance liquid chromatography. Our findings reveal that ancestors of yerba mate’s caffeine synthase enzymes were able to methylate a wide variety of substrates to form the beginnings of a caffeine biosynthetic pathway, but were not able to catalyze the final step to render caffeine. It wasn’t until three highly specialized modern caffeine synthase enzymes evolved following gene duplication that a full pathway could be complete. Furthermore, preliminary mutational analysis supports the idea that relatively few mutations within key regions of caffeine synthase genes are largely responsible for the transition to the highly specialized modern enzymes responsible for caffeine production found in yerba mate today. In summary, our work allowed us to better understand how yerba mate’s caffeine biosynthetic pathway arose and adds to the knowledge base about how multistep pathways form and render valuable plant metabolites, like caffeine.
Recommended Citation
Smith, Madeline, "Understanding the evolution of the genes and pathways responsible for caffeine production in Ilex paraguariensis" (2024). Honors Theses. 3849.
https://scholarworks.wmich.edu/honors_theses/3849
Access Setting
Honors Thesis-Restricted
Restricted to Campus until
6-19-2025