Unveiling Enzymatic Dynamics and Intermediates: Time-Resolved Serial Crystallography of Luciferase Catalysis with Innovative In-Situ Oxygen Delivery Techniques

• Applicant: Dr. Johannes Lang
• Subject Area: Structural Biology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 564012687

Enzymes are proteins that facilitate countless chemical reactions in the cells of every organism, making them essential for metabolism and biosyntheses. Enzymes that enable the oxidation of their substrate are called oxygenases and utilise oxygen for this reaction. A special subgroup of oxygenases are luciferases which are unique in producing a photon of light during their reaction. This phenomenon underlies the remarkable process of bioluminescence observed in organisms such as fungi, corals, and insects like fireflies. Scientists leverage this effect in biomedical basic research and drug development. Despite the extensive biomedical applications of luciferases, only little is known about the detailed reaction pathways and dynamic processes during the enzymatic conversion of their substrates. The goal of this project is to use cutting-edge time-resolved serial crystallography to elucidate these structures and processes at an atomic level – an endeavour that has yet to be achieved in the field of luciferases and related enzymes. Since luciferases are oxygen (O₂)-dependent, and the required O₂ concentrations in crystals can only be partially achieved with currently available methods, this project also aims to develop new techniques for rapid and sufficient O₂ delivery. This advancement would simplify the identification of intermediate structures. A particular focus is placed on enzymes that generate large amounts of O₂ upon substrate addition. Another promising approach involves photocages – chemical compounds that release O₂ upon laser excitation. Optimising these methods could significantly advance gas-dependent time-resolved serial crystallography and other spectroscopic techniques, benefiting the entire research community. The anticipated outcomes of this project will provide insights into the precise dynamic control of catalytic processes, deepening our understanding of enzymatic functions and opening new avenues for the targeted application and manipulation of these reactions.

• DFG Programme: WBP Fellowship
• International Connection: United Kingdom
• Host: Dr. Patrick Rabe