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Covering: 2013 to 2020The electron cryo-microscopy (cryo-EM) method Microcrystal Electron Diffraction (MicroED) allows the collection of high-resolution structural data from vanishingly small crystals that appear like amorphous powders or very fine needles. Since its debut in 2013, data collection and analysis schemes have been fine-tuned, and there are currently close to 100 structures determined by MicroED. Although originally developed to study proteins, MicroED is also very powerful for smaller systems, with some recent and very promising examples from the field of natural products. Herein, we review what has been achieved so far and provide examples of natural product structures, as well as demonstrate the expected future impact of MicroED to the field of natural product and small molecule research.

Microcrystal Electron Diffraction (MicroED) is the newest cryo-electron microscopy (cryo-EM) method, with over 70 protein, peptide, and several small organic molecule structures already determined. In MicroED, micro- or nanocrystalline samples in solution are deposited on electron microscopy grids and examined in a cryo-electron microscope, ideally under cryogenic conditions. Continuous rotation diffraction data are collected and then processed using conventional X-ray crystallography programs. The protocol outlined here details how to obtain and identify the nanocrystals, how to set up the microscope for screening and for MicroED data collection, and how to collect and process data to complete high-resolution structures. For well-behaving crystals with high-resolution diffraction in cryo-EM, the entire process can be achieved in less than an hour.