The many faces of radiation-induced changes

Abstract

During diffraction experiments even cryo-cooled protein crystals can be significantly damaged due to chemical and physical changes induced by absorbed X-ray photons. The character and scale of the observed effects depend strongly on the temperature and the composition of crystals. The absorption of radiation energy results in incremental regular changes to the crystal structure, making its impact on the process of solving the structure strongly correlated with other experimental variables. An understanding of all the dependencies is still limited and does not allow for a precise prediction of the outcome of a particular diffraction experiment. Results are presented of diffraction experiments performed under different experimental conditions. The influence of temperature and crystal composition on different characteristics of radiation damage is analyzed. The observed effects are discussed in terms of their impact on data processing and phasing procedures.

Diffraction data analysis in the presence of radiation damage

“…As noticed by Kmetko et al (2006) and also by others (Borek et al, 2007), accumulating X-ray dose at a particular temperature produces the same scaling B-factor increase irrespectively of how the same dose was generated by different combinations of mass-absorption coefficients, exposure time and beam intensity. This relationship between the dose and the B rel increase can be explained by the statistical properties of atom displacements generated by X-ray radiation.…”

“…For uncooled crystals, it is the dominant source of resolution-dependent intensity decay. However, for cryocooled crystals unit-cell changes are typically so minuscule (Borek et al, 2007;Ravelli et al, 2002;Murray & Garman, 2002) that the contribution of lattice disorder to the overall diffraction decay is not significant. In some special cases, discussed in x3.1.3, nonlinear effects may cause lattice disorder, but this will have more consequences than solely the overall decay of the diffraction pattern.…”

“…A data set from a crystal of bovine pancreatic trypsin inhibitor (BPTI) corresponding to PDB code 1g6x was provided by Mariusz Jaskolski and was collected on EMBL beamline X11 at DESY as described previously (Addlagatta et al, 2001). A data set from a crystal of NaI-841 (Borek et al, 2007), isomorphous with PDB deposition 1r61, was provided by Jana Maderova, who collected it on beamline 19-BM of the SBC at the APS. A data set from a crystal of thiamine-binding lipoprotein p37 from Mycoplasma hyorhinis, referred to here as p37n33 (C. Dann, unpublished work), was provided by Charles Dann III, who collected it at Cu K wavelength with an FR-E Super Bright X-ray generator (Rigaku).…”

“…The decay of half the total diffraction intensity defines the upper limit for X-ray dose (Henderson, 1995;Owen et al, 2006;Kmetko et al, 2006). However, chemical and physical changes induced by X-ray photons during data collection may result in the deterioration of merging statistics, sometimes long before reaching half the intensity decay (Borek et al, 2007;Zwart et al, 2004), prompting experimenters to limit the exposure. Even if limiting the exposure preserves less damaged states of the crystal, it often hinders solution of a structure owing to the collected data being either too weak or having insufficient multiplicity of observations to perform experimental phasing .…”

“…The identified patterns may be used at various points of different crystallographic procedures: (i) to plan the next experiment (sometimes even a continuation of the current one; Bourenkov & Popov, 2006); (ii) to correct data in reciprocal space, e.g. to apply a decay correction (Borek et al, 2007;Otwinowski et al, 2003) and zero-dose extrapolation (Borek et al, 2007;Diederichs et al, 2003;Blake & Phillips, 1962); and (iii) to combine real-space and reciprocal-space patterns for phasing Schiltz & Bricogne, 2007;Schiltz et al, 2004) In this article, we describe the main data patterns, how to recognize these, when to expect them and how they have impacted on data collection and phasing in a range of specific cases.…”