To reduce radiation damage of the radicals formed by ionizing X-ray photons, data collection is usually performed at ~100 K, prolonging significantly the life of the crystal (see Nave, C. & Garman, E.F. Towards an understanding of radiation damage in cryocooled macromolecular crystals. Journal of synchrotron radiation 12, 257–260 (2005): "up to 70 times more absorbed dose can be delivered to a cryocooled macromolecular crystal compared with one at room temperature before significant degradation in crystal order occurs").
Cryogenic temperatures (~100 K, or 90-120 K) close to the boiling point of liquid nitrogen (77 K) reduces thermal vibrations, and flexible parts of the protein can become distinct in electron density.
Symptoms of radiation damage:
During data collection
decrease in diffraction intensity
decrease in resolution
increase in unit cell volume
During model rebuilding-refinement
breakage of disulfide bonds
demethylation of Met
decarboxylation of Asp, Glu and the C-ter
loss of OH from Tyr
Ref. Biomolecular Crystallography By Bernhard Rupp
A review paper published in 2004,
Cryoprotectant database for protein crystals:
A snapshot from Garman and Schneider. Macromolecular cryocrystallography. J Appl Crystallogr (1997) vol. 30 (3) pp. 8898
A large number of compounds have been used successfully for the cryoprotection of macromolecular crystals (Table 1). [...] Finding the right cry-oprotectant for a particular system is to some extent a matter of trial and error. In many instances, the cryoprotection of crystals grown from aqueous/organic mixtures or from buffers of low salt concentration can be achieved easily by the addition of organic compounds [...] in a large proportion of these cases the following guidelines will lead to success. If a polyethylene glycol (PEG) with a molecular weight of less than 4000 or MPD is used as the precipitating agent, increasing the concentration of the precipitant can yield a good cryobuffer. For crystals induced by PEGs with molecular weights of 4000 and higher, the addition of small PEGs (i.e. PEG 400 or PEG 600) is recommended. For crystals grown by salting out at relatively low salt concentrations, MPD and ethylene glycol are promising candidates. In other cases, glycerol at concentrations up to 50% seems to be an almost universal cryoprotective agent for a wide range of commonly used crystallization buffers.
Pflugrath JW (2004) Macromolecular cryocrystallography--methods for cooling and mounting protein crystals at cryogenic temperatures. Methods 34:415–423.