Four-Dimensional Cryo Electron Microscopy at Quasi Atomic Resolution: "IMAGIC 4D"

The traditional tools of the structural biologist seeking to understand macromolecules and their complexes are X-ray crystallography and NMR spectroscopy. Single-particle cryo-electron microscopy (cryo-EM) has established itself as a new structural-biology technique over the last 15 years. Spectacular insights into the functioning of macromolecular complexes have been achieved especially from combining cryo-EM with the earlier approaches. The resolution levels achieved improved over the last decade from ∼10 A to sometimes better than ∼4 A, meaning that a de novo structure determination based on single-particle cryo-EM studies alone is now feasible. More challenging is the new perspective that cryo-EM brings: sorting heterogeneous populations of molecules into individual three-dimensional conformers resulting in sequences of related three-dimensional structures, or in short ‘4D cryo-EM’. Thanks to these developments, single-particle cryo-EM has become the technique of choice for shedding light on the functioning of many a complex biological system. The design of the software instrumentation for 4D cryo-EM is crucial. In this chapter we elaborate on organization issues for single-particle cryo-EM software, as exemplified by recent developments in the IMAGIC 4D software system. Keywords: cryo-electron microscopy; IMAGIC 4D