What Continuous evolution (CE) of proteins is an emerging laboratory technique that allows accelerated exploration and creation of protein function. In this project, variants of the CE technique will be established and used to: 1) predict the emergence of drug resistance in enzymes of natural pathogens, and 2) design functional non-natural proteins that bind small molecules (e.g. dyes). The first part will investigate the risk and mechanism of drug-induced mutation in the main protease of SARSCoV2 using non-hazardous and high-throughput CE experiments. The second part explores the frontier of de-novo protein sensor and actuator design with a focus on generating small molecule binding sites using deep learning algorithms combined with efficient experimental optimization via CE. Why With accelerated generation of protein mutants via CE, the project aims to characterize the evolutionary barriers of natural enzymes and refine methods for non-natural protein design. The gained biophysical and biochemical insight can advance: 1) drug development practices by fast identification of drug resistance mechanisms, and 2) protein-ligand complex engineering. As for the latter, remarkable progress has recently been made in the computational prediction and design of static protein structures. The next frontier is to apply these methods to design functional proteins. To this end, exploring protein plasticity as well as specific interactions with ligands are critical as functional proteins are often characterized by flexibility, reactivity, and small-molecule binding. How The project and the future envisioned program are highly interdisciplinary with origins in chemical biology and biophysical chemistry. The Reintegration Fellowship will allow me to return from Stanford University to the Department of Chemistry at Aarhus University. The department has a strong existing infrastructure and workshop expertise that will be beneficial for developing new experimental setups for efficient directed protein evolution. The project naturally intersects with expertise at the department in small molecule synthesis and molecular modelling. Overall, the strong interdisciplinary environment at Aarhus University within molecular biology, nanoscience and biomedicine offers a dynamic environment for this project to succeed and flourish into an independent research program. SSR To curtail the current and future coronavirus pandemics, the project explores the evolutionary barriers of the coronavirus main protease, a key drug target. Various protease inhibitors have already been reported including the promising PF-007321332. Accelerated and non-hazardous methods for accessing drug resistance risk could help guide prioritization of promising drug candidates early in the development process. Long-term, the project aims to establish synergies between computational and experimental methods that will enhance our abilities to design and discover functional protein-ligand complexes. Examples include novel light harvesting and converting complexes with relevance for green chemistry, as well as protein switches and sensors for cell- and protein-based molecular medicine.