Abstract
Serial X-ray crystallography at free-electron lasers allows to solve biomolecular structures from sub-micron-sized crystals. However, beam time at these facilities is scarce, and involved sample delivery techniques are required. On the other hand, rotation electron diffraction (MicroED) has shown great potential as an alternative means for protein nano-crystallography. Here, we present a method for serial electron diffraction of protein nanocrystals combining the benefits of both approaches. In a scanning transmission electron microscope, crystals randomly dispersed on a sample grid are automatically mapped, and a diffraction pattern at fixed orientation is recorded from each at a high acquisition rate. Dose fractionation ensures minimal radiation damage effects. We demonstrate the method by solving the structure of granulovirus occlusion bodies and lysozyme to resolutions of 1.55 Å and 1.80 Å, respectively. Our method promises to provide rapid structure determination for many classes of materials with minimal sample consumption, using readily available instrumentation.
Highlights
Serial X-ray crystallography at free-electron lasers allows to solve biomolecular structures from sub-micron-sized crystals
We perform protein crystallography by SerialED using a parallel nanobeam in a scanning transmission electron microscope (S/TEM)
Our results show that SerialED allows the determination of protein structures at high resolution from extremely small protein crystals in a rapid, efficient, and automated manner
Summary
Serial X-ray crystallography at free-electron lasers allows to solve biomolecular structures from sub-micron-sized crystals. While seminal experiments on 2D crystals[20] were restricted to a small class of suitable samples, various successful implementations of 3D rotation electron diffraction (3D ED) solving structures of beam-sensitive small molecules[21,22,23] sparked interest in applying 3D crystallography to biomolecules, a technique referred to as MicroED24–26. We apply SerialED to protein nanocrystals, using a doseefficient automated data collection scheme that enabled us to solve the highest-resolution protein structure by ED to date This method provides a viable alternative to serial femtosecond crystallography for the determination of high-resolution protein structures from sub-micron-sized crystals using laboratory-based instrumentation
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