Drop on fixed target reaction initiation approach for serial and time resolved crystallography

Abstract

We describe the design and implementation of a drop on fixed target method for time-resolved serial crystallography at both synchrotron and XFEL facilities. A piezoelectric droplet dispensing pipette is employed for addition of picolitre volume (40 - 90 pL) aqueous droplets, containing (co-)substrate(s) or ligand(s), onto enzyme microcrystals immobilised on a solid support. The system was tested with various enzyme systems, including lysozyme and two -lactamases, CTX-M-15 and AmpCEC. Mitigation strategies for cross-well contamination, including the implementation of interleaved controls, are described; the overall performance of the system at synchrotron and X-ray free electron laser facilities was evaluated. This drop on fixed target method is a reliable framework for time-resolved crystallography and will improve the consistency of measurements across facilities.