Facing the phase problem

The marvel of X-ray crystallography is the beauty and precision atomic structures deduced from diffraction patterns. Since these patterns record only amplitudes, phases for diffracted waves must also be evaluated systematic structure determination. Thus, we have phase problem as a central complication, both intellectually field practically so many analyses. Here, I discuss how – myself, my laboratory community faced problem, considering evolution methods evaluation structural biology developed to present day. During explosive growth macromolecular crystallography, practice in analysis evolved universal reliance on isomorphous replacement eventual domination anomalous de novo As Protein Data Bank (PDB) grew familial relationships among proteins became clear, molecular overtook all other phasing methods; however, experimental remained essential molecules without obvious precedents, with multi- single-wavelength (MAD SAD) predominating. While mathematics-based direct had proved inadequate typical macromolecules, they returned crack substantial selenium substructures SAD analyses selenomethionyl proteins. Native SAD, exploiting intrinsic S P atoms biomolecules, has become routine. Selenomethionyl MAD were mainstays genomics efforts populate PDB novel A recent dividend been paid success PDB-trained artificial intelligence approaches protein prediction. Currently, AlphaFold models often obviates need evaluation. For multiple reasons, are now unfazed by problem. Cryo-EM an attractive alternative applications today's biologists. It simply finesses problem; principles procedures remain pertinent adopted single-particle cryo-EM studies biomolecules.