The temperature of flash-cooling has dramatic effects on the diffraction quality of nucleosome crystals

Abstract

Nucleosome core-particle crystals are routinely flash-cooled in liquid propane at temperatures of approximately 153 K, followed by transfer into a cold nitrogen-gas stream (approximately 93 K). Analysis of diffraction data from crystals flash-cooled at different temperatures shows that the optimal temperature is approximately 153 K. The data quality worsens, with a concomitant reduction in the diffraction limit, at temperatures both higher and lower than 153 K. With some batches of crystals, significant shrinkage of the unit-cell volume is also observed at temperatures of 138 K and lower. The lattice shrinkage is always restricted to the c axis, concurrent with closer packing of two nucleosomes. Direct plunge-cooling of crystals in liquid nitrogen leads to loss of diffraction quality and resolution limit. Thus, in cases where flash-cooling into liquid nitrogen is detrimental to diffraction, optimizing cooling protocols at higher temperatures using liquid propane or other cryogens with similar properties may lead to dramatically improved results. In a related study, it is shown that a nucleosome crystal transported under 'cryocooled' conditions has higher mosaicity and yields inferior data quality in comparison to a crystal cryocooled at the synchrotron. For fragile crystals, transport in mother liquor and/or cryoprotectant followed by subsequent flash-cooling at the synchrotron may be the best procedure.