Serial box shot 3d 3.6 serial number series#

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 also to biomolecules, a technique also referred to as MicroED 24, 25, 26. Compared to X-rays, the obtainable diffraction signal for a given crystal volume and tolerable radiation dose is up to three orders of magnitude larger due to the higher ratio of elastic to inelastic electron scattering events and a much smaller energy deposition per inelastic event 1, 19. While the low penetration depth of electrons renders them unsuitable for large three-dimensional crystals, their physical scattering properties are specifically advantageous for sub-micron crystals of radiation-sensitive materials. The development of serial crystallography using smaller scale, ideally laboratory-based instrumentation is therefore highly desirable.Įlectron microscopes are a comparatively ubiquitous and cost-effective alternative for measuring diffraction from nanocrystals. However, the scarcity and costliness of XFEL beamtime limits the use of protein nanocrystals for routine structure determination.

Ideally, radiation-damage effects are entirely evaded either by a “diffract-before-destroy” mode using femtosecond XFEL pulses 5 or by imposing doses too low to cause significant structural damage of each crystal, which has also been implemented at synchrotron micro-focus beam lines 11, 12, 18. Sufficient signal-to-noise ratio and completeness is achieved through merging of many thousands of such snapshots.

Serial box shot 3d 3.6 serial number series#

Here, acquiring snapshots in a single orientation from each crystal instead of a rotation series avoids dose accumulation, permitting higher fluences, which concomitantly decreases the required diffracting volume. Notably, X-ray free-electron lasers (XFELs) have driven the development of serial crystallography 5, 6, 7, 8, 9, 10, a technique that is also increasingly applied at synchrotron sources 11, 12, 13, 14, 15, 16, 17. However, during the past few years crystallographic techniques have emerged that are able to exploit nanocrystals for diffraction experiments. Sub-micron crystals can be obtained more readily and are a common natural phenomenon, but often escape structure determination as the small diffracting volume and low tolerated dose of typically tens of MGy 3, 4 prohibit the measurement of sufficient signal. An important limitation of biomolecular crystallography lies in the difficulty to obtain large, well-ordered crystals, which is particularly prevalent for membrane proteins and macromolecular complexes. This includes the majority of membrane proteins, which are often too small for computational alignment as required by single-particle analysis 1, 2. Despite recent advances in single-particle cryo-electron microscopy (cryo-EM), the vast majority of high-resolution structures are determined by crystallographic methods ( ). Our method promises to provide rapid structure determination for many classes of materials with minimal sample consumption, using readily available instrumentation.Īn understanding of macromolecular structure is crucial for insight into the function of complex biological systems. 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.

Dose fractionation ensures minimal radiation damage effects. 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. Here, we present a method for serial electron diffraction of protein nanocrystals combining the benefits of both approaches.

On the other hand, rotation electron diffraction (MicroED) has shown great potential as an alternative means for protein nano-crystallography. However, beam time at these facilities is scarce, and involved sample delivery techniques are required. Serial X-ray crystallography at free-electron lasers allows to solve biomolecular structures from sub-micron-sized crystals.