As XFELS grow out of their infancy and begin to proliferate, the techniques and methodology mature. While work at synchrotrons and other X-ray light sources will continue to contribute heavily towards the growing knowledge base of macromolecular structures, investigating dynamics using these instruments remains highly restricted. Through serial femtosecond crystallography (SFX), XFELs offer the ability to probe structural dynamics both in the ultrafast regime and at a much finer time scale, pushing towards the realization of true “molecular movies” where conformational and enzymatic dynamics can be understood in the time regimes that they actually occur.
This advantage is underscored by the ultra-compact femtosecond pulses and nano- and micron sized crystals utilized in SFX. The ultrafast time regime allows higher temporal resolution along a reaction timeline as well as the ability to probe much shorter delay times than can be realized with other structural techniques. The exploitation of small crystals compliments this, allowing a tighter temporal reaction homogeneity which is particularly important for short lived intermediates. Many pump-probe style experiments have been successfully achieved using XFELs to date, extending from validating proof-of-principle and boundary establishing techniques to novel understanding of macromolecular dynamics, comprising the first true “molecular movies”. More recently, diffusion based “on-the-fly” mixing experiments have been successfully conducted, paving the way for a whole new level of understanding in enzymatic structure-function relationships. As structural biology at XFELs emerges as an established technique, time-resolved studies exemplify the incredible potential of what this field can offer to the scientific community at large.