Crystallogenesis studies in microgravity with the Advanced Protein Crystallization Facility on SpaceHab-01

Abstract

The Advanced Protein Crystallization Facility (APCF), a new protein crystallization device developed by ESA for the IML-2 Mission in 1994, was tested in its maiden flight on STS-57 Mission in SpaceHab-01 with a physico-chemical experiment on lysozyme crystallization. In pre-flight ground experiments, prior to the Shuttle Mission, the protocol for lysozyme crystallization with NaCl was based on its solubility diagram at 18°C and pH 4.5. Crystallization was conducted under microgravity in 25 APCF reactors using vapor diffusion, dialysis, and free liquid interface diffusion, with control on earth in 25 identical reactors. Identical supersaturation values were tested by the three crystallization techniques. Values of supersaturation derived from ground experiments allowed for conditions that yielded crystals in microgravity. The average number and size of crystals from the flight experiment and the earth control showed no significant difference; however many crystals were not free floating and grew on the walls of some of the protein chambers. The dialysis technique proved to be suitable, since no additional nucleation was generated by the membrane. Protein concentration measurements indicated that 13 days after activation of the experiment as much as 70–90% of the protein in supersaturated state had already crystallized. Data indicated differences in the crystallization behavior depending upon the crystallization set-up. Images of the protein chamber of 6 reactors, recorded during the flight, allowed us to evaluate the early stage of crystallization, to verify that recovered crystals had actually grown under microgravity conditions, and showed motions of crystals during the Mission. Using synchrotron radiation, resolution and rocking curve measurements of ground and space lysozyme crystals grown in APCF reactors showed no significant differences, although the values are much better than previously recorded diffraction limits and mosaicity data obtained with tetragonal lysozyme crystals grown in other set-ups and under different conditions. All controls foreseen throughout the microgravity experiment proved to be essential for the interpretation of the flight data, as concerning the effect of microgravity.