Similarities and differences in radiation damage at 100 K versus 160 K in a crystal of thermolysin.

The temperature-dependence of radiation damage in macromolecular X-ray crystallography is currently much debated. Most protein crystallographic studies are based on data collected at 100 K. Data collection at temperatures below 100 K has been proposed to reduce radiation damage and above 100 K to be useful for kinetic crystallography that is aimed at the generation and trapping of protein intermediate states. Here the global and specific synchrotron-radiation sensitivity of crystalline thermolysin at 100 and 160 K are compared. Both types of damage are higher at 160 K than at 100 K. At 160 K more residue types are affected (Lys, Asp, Gln, Pro, Thr, Met, Asn) than at 100 K (Met, Asp, Glu, Lys). The X-ray-induced relative atomic B-factor increase is shown to correlate with the proximity of the atom to the nearest solvent channel at 160 K. Two models may explain the observed correlation: either an increase in static disorder or an increased attack of hydroxyl radicals from the solvent area of the crystal.