Protein-to-structure pipeline for ambienttemperature in situ crystallography at VMXi

The utility of X-ray crystal structures determined under ambient-temperature conditions is becoming increasingly recognized. Such experiments can allow protein dynamics to be characterized and are particularly well suited to challenging protein targets that may form fragile crystals that are difficult to cryo-cool. Room-temperature data collection also enables time-resolved experiments. In contrast to the high-throughput highly automated pipelines for determination of structures at cryogenic temperatures widely available at synchrotron beamlines, room-temperature methodology is less mature. Here, the current status of the fully automated ambient-temperature beamline VMXi at Diamond Light Source is described, and a highly efficient pipeline from protein sample to final multi-crystal data analysis and structure determination is shown. The capability of the pipeline is illustrated using a range of user case studies representing different challenges, and from high and lower symmetry space groups and varied crystal sizes. It is also demonstrated that very rapid structure determination from crystals in situ within crystallization plates is now routine with minimal user intervention.

Automated summary

The utility of X-ray crystal structures at ambient-temperature conditions is increasingly recognized due to their ability to characterize protein dynamics and address challenging protein targets. The fully automated ambient-temperature beamline VMXi at Diamond Light Source provides an efficient pipeline for protein sample to final structure determination, as demonstrated through various user case studies. It is also shown that rapid in situ structure determination from crystals within crystallization plates is now routine with minimal user intervention.