Refinement of amino-acid conformation vs. difference density maps in time-resolved serial femtosecond crystallography...

Time-resolved crystallography is a revolutionary X-ray diffraction technique by which the structural features of short-lived, transient intermediates of in crystallo reactions can be elucidated. While visualizing time-dependent structural changes via difference electron density maps is relatively simple, time-resolved diffraction data is complex because it arises from a substrate-dominated mix of the different reaction components. Thus, atomic coordinate refinement of intermediate species is challenging and prone to bias, as it requires deconvolution of the mixed-states. To simplify the refinement process, we have developed difference electron density correlation coefficient real space refinement (dFoCC refinement). By basing coordinate refinement on comparing observed vs. calculated difference density maps, dFoCC produces reasonable atomic coordinates of intermediate species in a reproducible manner and with clearly defined quality metrics.