The high-mosaicity illusion: revealing the true physical characteristics of macromolecular crystals

Typical measurements of macromolecular crystal mosaicity are dominated by t he characteristics of the X-ray beam and as a result the mosaicity value gi ven during data processing can be an artifact of the instrumentation rather than the sample. For physical characterization of crystals, an experimenta l system and software have been developed to simultaneously measure the dif fraction resolution and mosaic spread of macromolecular crystals. The contr ibutions of the X-ray beam to the reflection angular widths were minimized by using a highly parallel, highly monochromatic synchrotron source. Hundre ds of reflection profiles over a wide resolution range were rapidly measure d using a charge-coupled device (CCD) area detector in combination with sup erfine phi-slicing data collection. The Lorentz effect and beam contributio ns were evaluated and deconvoluted from the recorded data. Data collection and processing is described. From 1 degrees of superfine phi-slice data col lected on a crystal of manganese superoxide dismutase, the mosaicities of 2 60 reflections were measured. The average mosaicity was 0.0101 degrees (s.d . 0.0035 degrees) measured as the full-width at half-maximum (FWHM) and ran ged from 0.0011 to 0.0188 degrees. Each reflection profile was individually fitted with two Gaussian profiles, with the first Gaussian contributing 55 % (s.d. 9%) and the second contributing 35% (s.d. 9%) of the reflection. On average, the deconvoluted width of the first Gaussian was 0.0054 degrees ( s.d. 0.0015 degrees) and the second was 0.0061 degrees (s.d. 0.0023 degrees ). The mosaicity of the crystal was anisotropic, with FWHM values of 0.0068 , 0.0140 and 0.0046 degrees along the a, b and c axes, respectively. The an isotropic mosaicity analysis indicates that the crystal is most perfect in the direction that corresponds to the favored growth direction of the cryst al.