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Technology Topics Crystallography

Optimizing Damage

SBKB [doi:10.1038/sbkb.2012.186]
Technical Highlight - February 2014
Short description: Radiation damage is inherent to the X-ray diffraction experiment. A new metric predicts crystal lifetime, allowing for more efficient and effective data collection strategies.

Dose contour maps of two contrasting diffraction data collection strategies, simulated using RADDOSE-3D. Blue to red represents low to high dose. Figure courtesy of Elspeth Garman.

Macromolecular crystallography uses the diffraction obtained from scattered X-rays to determine the structure of the protein molecules that make up the crystal. Radiation damage to the crystal is an unavoidable part of the diffraction experiment, leading to decay in diffraction intensity and an increase in the relative B factor over time. It is one of the most frequent causes of failure in crystallography experiments.

Researchers can reduce radiation damage by lowering the number of X-rays, but this also leads to weaker diffraction; thus, a balance must be struck between diffraction intensity and crystal damage. Particularly in the case of more challenging crystallographic experiments, where there are only a limited number of well-diffracting crystals, the ability to predict the “effective crystal lifetime” that would be available to perform a diffraction experiment most efficiently is important. Two available metrics are the average energy absorbed per unit mass (or dose) for the whole crystal volume (which can be calculated by the program RADDOSE-3D) and the maximum dose, which is the highest dose reached at any point in the crystal. However, both of these metrics only report on the global dose state at the end of the diffraction experiment.

Now, Garman and colleagues have derived a diffraction-weighted dose (DWD) metric that can provide a time- and space-resolved accounting of the dose state of the crystal at any point during the diffraction experiment. In short, the DWD provides the average dose for the crystal volume that is contributing to a given diffraction pattern. Using data collected from cryocooled bovine pancreatic insulin crystals irradiated in X-ray beams of three different sizes, two of which resulted in uneven dose profiles since the beams were smaller than the crystals, the authors found that the DWD was able to accurately predict intensity decay under all three beam conditions, and so it could be used to predict crystal lifetime.

Further studies showed that DWD can be used to optimize the geometry of the diffraction experiment by maximizing the diffracted dose efficiency (DDE). Typical diffraction experiments have the beam and rotation axes well-aligned, so the central point of maximum dose is where these two axes intersect. In an offset approach, the rotation axis is shifted and the maximum dose region is a torus around the rotation axis. This results in a lower peak dose. The authors found that, depending upon the size of the crystal and beam being used, the DDE can be improved if an offset geometry is used. In one test case, the use of an offset strategy allowed for 30% longer overall exposure time and a 25% higher diffraction yield for the same diffraction intensity decay.

In combination, DWD and DDE can be used to predict crystal lifetime and optimize data collection strategies. While the authors concede that they were using morphologically optimized crystals, online three-dimensional imaging of the crystal and two-dimensional imaging of the beam are technologies that are currently being integrated into some beamlines, which would allow for the routine implementation of DWD.

Michelle Montoya


  1. O.B. Zeldin et al. Predicting the X-ray lifetime of protein crystals.
    Proc Natl Acad Sci U S A. 110, 20551-6 (2013). doi:10.1073/pnas.1315879110

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