PIF2024 - NUEVAS APROXIMACIONES EN CRISTALOGRAFIA RESUELTA EN EL TIEMPO EN LA ESCALA DE MILISEGUNDOS PARA ESTUDIOS ESTRUCTURALES Y DINAMICOS EN 3D DE ENZIMAS DE INTERES TERAPEUTICO (PID2023-151100NB-I00)

Enzymes drive life by catalyzing nearly all biochemical reactions, including metabolic processes, energy transfer, genetic information processing, and signaling. Despite extensive research, a key question remains: how do structural fluctuations in enzymes enhance their ability to catalyze complex reactions? Understanding this could revolutionize biology and biotechnology, leading to insights into enzyme efficiency, novel enzyme activities, and the design of artificial biocatalysts for therapeutic and industrial use. While X-ray crystallography has traditionally been static, advances in time-resolved serial crystallography now allow us to observe biomolecular reactions and structural changes with near-atomic resolution. In this project we will use time-resolved serial femtosecond crystallography at X-ray free-electron lasers and synchrotrons to determine 3D structures of transient intermediates with ultrafast (100 fs) time resolution with the aim at establishing the timescales of structural changes in enzyme crystals compared to those in solution. Our novel approach involves initiating a chemical reaction in protein nanocrystals and tracking its progress with time-resolved serial crystallography. We will identify key time points to determine the 3D structures of intermediates and use hybrid quantum mechanics/molecular mechanics and molecular dynamics simulations to validate our data. This will provide unprecedented insights into the reaction mechanisms of enzymes such as Anmk.