Time-resolved SSX: methodologies and applications to enzymes and light-sensitive proteins

Time-resolved serial synchrotron crystallography (TR-SSX) poses methodological challenges, yet offers novel opportunities to study structural changes of biological macromolecules at work in real time. Our proposal addresses the former and presents various examples to grasp the latter. We will implement microfluidic, jet-based and solid-support sample delivery methods and combine their use with different means of reaction triggering in the crystalline molecules including rapid mixing and light-illumination. Studies will be conducted to assess specific and global X-ray radiation damage at the unprecedented flux densities ID29 will deliver. TR-SSX will then be applied to several enzymes and light-sensitive proteins to study structural changes on the micro- to millisecond time-scale following reaction triggering. TR-SSX and XFEL based TR-SFX will be compared by conducting a study on the micro- to millisecond time-scale on the same protein crystals delivered to the beam by the same method.