Electron diffraction captures high-resolution structures from in vivo protein nanocrystals of Bacillus thuringiensis

Bacillus thuringiensis is one of the most widely used biopesticides worldwide owing to the highly specific pesticidal-proteins various strains produce in the form of nanocrystals. Structure determination of such crystals remains difficult because their small size makes them unsuitable for conventional X-ray crystallography. Here we explore two emerging (cryo-) electron diffraction techniques, namely three-dimensional electron diffraction and serial electron diffraction, as tools for studying the structures of these crystals. Using the mosquitocidal protein Cry11Aa as an example, we compare electron diffraction with state of the art results obtained with an X-ray free electron laser. Our work demonstrates that electron diffraction is a viable alternative for structure determination from such challenging crystals, in some cases outperforming previous results obtained with X-ray free electron lasers. We present a workflow based on readily available instrumentation enabling structure determination directly from the crystals grown in vivo, unperturbed by dissolution and therefore preserved in their native state.