Setting high-resolution limits for anisotropic data
Melanie Vollmar, G. N. Murshudov, F. von Delft · Acta Crystallographica Section A Foundations of Crystallography · 2012
Most diffraction data are anisotropic and often strongly so, which can significantly affect the ability to interpret derived electron density, making model building and structure refinement very challenging.There is no consensus on how anisotropy should be treated at the resolution limit: imposing isotropic resolution limits either excludes many reflections with good signal, or includes reflections without signal.The problem has various approaches both at data integration and structure refinement.Data integration with HKL2000 effectively imposes anisotropic resolution limits, while MOSFLM/SCALA and XDS both apply isotropic resolution limits, yet generate significantly differing estimates of standard deviations of intensitiesin regions where diffraction is weak.These standard deviationsnot only inform subsequent refinement, but the various programs treat these estimates very differently, leading to very different electron density.A popular variation is to modify measured amplitudes using the Diffraction Anisotropy Server in an attempt to compensate for the anisotropy (http://services.mbi.ucla.edu/anisoscale/).We thus systematically examined the various approaches using a series of strongly anisotropic datasets with resolution below 2.6A.By assessing the residual density after refinement in REFMAC5, we compare the overall outcome of refinement and structure completeness when applying anisotropic, conservative isotropic and generous isotropic resolution cutoffs.In addition, we assess the use of different estimates of standard deviation from various integration/scaling packages, as well as anisotropic B-factor sharpening applied both by the Diffraction Anisotropy Server and within REFMAC5.Finally, standard deviationsare treated in a variety of ways during refinement, including use of intensities rather than amplitudes.A comparative analysis of the different approaches will be presented.