This thesis explores how to design and study enzyme reaction networks (ERNs) inside flow reactors, systems where liquids continuously move through a device to carry out biochemical reactions. It identifies the key principles needed to make these networks perform efficiently and to extract meaningful information from them.
It first examines how enzymes can be attached to tiny beads so they remain active under flow conditions. Building a multi enzyme pathway this way is challenging, as all enzymes must stay functional and work together smoothly. The thesis also evaluates different reactor types to determine which best support immobilised enzymes.
A major focus is on active learning employing experimental design approach that uses carefully varied input patterns to reveal how enzymes behave and interact. When enzymes are used freely in solution rather than attached to beads, this method becomes even more powerful, enabling faster discovery of reaction behaviour and reducing the number of experiments required.
The core finding is that timed input strategies, precisely controlling when and how substrates enter the system, can dramatically improve the final product yield of complex enzyme cascades. The thesis concludes by showing how these insights accelerate progress in cell free biosynthesis, a promising route to sustainable chemical production.
Miglė Jakštaitė was born on September 13, 1994, in Šiauliai, Lithuania. She enrolled at Vilnius University, Lithuania, where she earned her BSc in Chemistry in 2017. Motivated by her passion of interaction between Chemistry and Biology, Miglė decided to move to the Netherlands and pursue a MSc degree in Chemistry with a specialisation in Chemistry for Life at Radboud University, Nijmegen. She joined the group of Prof. Wilhelm Huck at the Department of Physical Organic Chemistry for her first internship, where she was constructing positive and negative feedback loops in enzymatic chemical reaction networks. After obtaining her MSc degree in Chemistry in 2019, Miglė chose to continue her academic journey and re-joined the group as a PhD candidate under the supervision of Prof. Wilhelm Huck at Radboud University. She researched dynamic control and optimisation of enzymatic reaction networks for cell free biosynthesis, the results of which are presented in this thesis.