Limiting the Effects of Radiation Damage in MicroED through Dose Selection during Data Processing

• Microcrystal electron diffraction (MicroED), also known as three-dimensional electron diffraction (3D ED), allows collection of diffraction data from submicron-sized crystals under low electron dose conditions, typically around 5-6 e-Å-2 in total. Despite having several advantages of MicroED over most conventional X-ray crystallographic techniques, susceptibility to radiation damage is a big problem that remains to be solved. Similar to X-ray crystallography, radiation damage to the macromolecular crystal structures in MicroED manifests in two forms, the global damage that affects the overall crystal lattice order and the site-specific damage that affects highly sensitive residues and moieties in macromolecules. In this study, we investigated data processing strategies that could be used to limit the effects of radiation damage to the crystal even when data collection is performed at high electron doses. During MicroED data collection, radiation damage increases with the number of acquired ED frames because the accumulated electron dose increases. To limit the damage, we propose to process only the first few frames of a dataset with a certain low dose cutoff. Data collected from several crystals and processed in this way can be merged to increase completeness and subsequently be used for structure refinement. According to our results, this approach improves the resolution of the data, the data statistics, the structure determination, and the quality of the final structure. The suggested approach could be especially useful in MicroED structure-based drug discovery where atomic resolution structures will provide detailed information about ligand-protein binding properties, which are essential during library screening and hit identification. 

Ämnesord

NATURVETENSKAP  -- Biologi -- Strukturbiologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Structural Biology (hsv//eng)

NATURVETENSKAP  -- Kemi -- Fysikalisk kemi (hsv//swe)

NATURAL SCIENCES  -- Chemical Sciences -- Physical Chemistry (hsv//eng)

Nyckelord

Microcrystal Electron Diffraction (MicroED)

Data processing

macromolecular structure

global radiation damage

site-specific radiation damage

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