Recent developments in structural biology, most notably the improvement in the structure determination by single particle cryo- electron microscopy (cryo-EM), the availability of X-ray free electron lasers (XFELs) and diffraction-limited synchrotron light sources for X-ray crystallography have moved the bottleneck in high-resolution structure determination of large biological assemblies to the biochemical preparation of high-quality samples. However, precisely the biochemical purification of such large, labile assemblies remains a formidable challenge and often fails when strategies suitable for single biomolecules are adapted to larger complexes, due to the dissociation of such samples resulting in structural heterogeneity.
Here, I will present strategies to purify and characterize high-quality samples of large biological assemblies. Benefitting from the strong complementarity of cryo-EM and X-ray crystallography, I will present structural biochemistry of proteasomes and fatty acid synthases at unprecedented resolution, new mechanistic findings and how these have opened up new venues for drug discovery. I will also discuss shortcomings of cryo-EM and present developments in synchrotron beamlines for such projects.