Dynamic response of tetragonal lysozyme crystals to changes in relative humidity: implications for post-growth crystal treatments

Abstract

The dynamic response of tetragonal lysozyme crystals to dehydration has been characterized in situ using a combination of X-ray topography, high-resolution diffraction lineshape measurements and conventional crystallographic diffraction. For dehydration from 98% relative humidity (r.h.) to above 89%, mosaicity and diffraction resolution show little change and X-ray topographs remain featureless. Lattice constants decrease rapidly but the lattice-constant distribution within the crystal remains very narrow, indicating that water concentration gradients remain very small. Near 88% r.h., the c-axis lattice parameter decreases abruptly, the steady-state mosaicity and diffraction resolution degrade sharply and topographs develop extensive contrast. This transformation exhibits metastability and hysteresis. At ®xed r.h. < 88% it is irreversible, but the original order can be almost completely restored by rehydration. These results suggest that this transformation is a ®rst-order structural transition involving an abrupt loss of crystal water. The front between transformed and untransformed regions may propagate inward from the crystal surface and the resulting stresses along the front may degrade mosaicity. Differences in crystal size, shape and initial perfection may produce the observed variations in degradation timescale. Consequently, the success of more general post-growth treatments may often involve identifying procedures that either avoid lattice transitions, minimize disorder created during such transitions or maintain the lattice in an ordered metastable state.

Slow cooling of protein crystals

“…Dehydration decreases the unit-cell volume and usually has little effect on mosaicity except at large dehydrations (Dobrianov et al, 2001). No evidence of dehydrationinduced cell-volume changes were observed in our control experiment, or for a sample held for 1 h at T = 300 K before irradiation.…”

Flash-cooling and annealing of protein crystals

“…X-ray topography measurements (Tanner, 1976;Fourme et al, 1995;Izumi et al, 1996;Dobrianov et al, 1998Dobrianov et al, , 1999Dobrianov et al, , 2001Otalora et al, 1999;Vidal et al, 1999) were performed on station F-3 by illuminating the entire crystal using a highly parallel (unfocused) monochromatic X-ray beam and recording the diffraction pattern using ®ne-grain high spatial resolution ®lm (Kodak Industrex SR) placed $5 cm from the sample. Under these illumination and detection conditions, every Bragg spot is a unit magni®-cation image of the crystal.…”

Plastic-embedded protein crystals

“…Several studies have been reported on the dynamic response of tetragonal lysozyme crystals to changes in relative humidity. For example, Dobrianov et al (2001) showed an irreversible unit-cell volume decrease of 9% upon changing the relative humidity around the crystal from 98% to 86%, while Deshpande et al (2005) showed that dehydratation by organic solvents mainly affected the order of the hydrated waters without causing large changes in the unit-cell dimensions. In our control study, where a lysozyme crystal was allowed to dehydrate in air for 15 h, we measured (100 K) a unit cell of a = 77.43 and c = 37.48 Å , which corresponds to a solvent content of 37%.…”