The application of enzymes in industry and medicine is dramatically expanding. However, the success of such applications depends on the ability to optimize enzyme properties to suit specific technological requirements. Nonetheless, to manipulate proteins effectively, their fundamental structures and functions must first be elucidated. In other words, the ability to engineer specific enzymes relies on understanding the mechanisms of enzyme catalysis. Conventional methods for investigating enzymatic reactions’ mechanistic properties are usually very complicated and demanding in terms of time and volume of reagents.
The international team developed a new laboratory method for analyzing enzyme reactions on a microfluidic chip, which performs a tremendous number of experiments in a very short time. The experiments transpire in tiny droplets of up to 5 picolitres, a million times smaller than a teardrop, with up to 9,000 reactions performed per minute. This allows performing about 200 million experiments from a single milliliter of reagent. In comparison, conventional micro-volume stopped-flow equipment would require over twenty 100 liter bathtubs of reagent for the same number of reactions, making the system at least of an order of magnitude better than the state-of-the-art equipment.
The on-chip measurements combined with modern mathematical data analysis methods and molecular modeling provide a deep understanding of enzymes, their behavior, and properties. “When we first applied the method described in the article, we observed an unusual phenomenon driving an enzymatic reaction with such detail, which would not be possible to obtain using conventional methods. This information shifts the understanding of the enzyme reaction principle, which helps us design new enzymes efficiently. We also see the on-chip technology's large potential for acquiring complex datasets for training artificial intelligence tools for medical and biotech research,” explains Zbyněk Prokop, who was responsible for developing the visualization methods for the enzyme reactions and analyzing the complex datasets collected from the on-chip experiments.
The project was successful due to the effective collaboration among many scientific disciplines. The scientists utilized the latest knowledge in microfluidics, microengineering, material chemistry, optics, biochemistry, mathematics, and computational science. The chip and optics' design and construction were carried out by Prof. Andrew deMello’s group, led by Dr. Stavros Stavrakis (ETH Zurich). For constructing the microfluidic chip, the team used soft lithography technology, a process taken from electronics for manufacturing integrated circuits. The computational simulations complementing the experimental observations were done by Prof. Jiri Damborsky and Dr. David Bednar’s team (The International Clinical Research Center, St. Anne's University Hospital, Brno). “An important component of this technology's successful development was the fantastic collaboration, excellent communication between the teams and resilience to multiple technical problems,” notes Dr. Stavros Stavrakis on the importance of this international scientific collaboration.
“The pressure for speed in research and development requires implementing a new generation of bioanalytical methods. Microfluidics offers a revolutionary solution, miniaturizing and integrating laboratory processes on silicon-based chips, in which thousands of reactions occur every second in miniature droplets. This technology has significantly changed the nucleic acid analysis field, allowing next-generation sequencing and modern diagnostics using digital PCR. We expect similar progress also in protein science,” concludes Prof. Zbyněk Prokop. The researchers aspire further to link this chip technology with advanced biophysical methods and for training artificial intelligence.
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