Comparative Analysis of Thaumatin Crystals Grown on Earth and in Microgravity

Abstract

The protein thaumatin was studied as a model macromolecule for crystallization in microgravity-environment experiments conducted on two US Space Shuttle missions (USML-2 and LMS). In this investigation, we have evaluated and compared the quality of space-and earth-grown thaumatin crystals using X-ray diffraction analyses, and characterized them according to crystal size, diffraction resolution limit and mosaicity. Two different approaches for growing thaumatin crystals in the microgravity environment, dialysis and liquid-liquid diffusion, were employed as a joint experiment by our two investigative teams. Thaumatin crystals grown in a microgravity environment were generally larger in volume and the total number of crystals was less, relative to crystals grown on earth. They diffracted to significantly higher resolution and with improved diffraction (/,,)

Investigating the Effect of Impurities on Macromolecule Crystal Growth in Microgravity

“…Mosaicity is due to a combination of three causes: 38,39 angular misalignment of domains within the crystal, finite domain volume, and lattice strain. In previous microgravity experiments, the reduced crystal mosaicity reported for lysozyme 4 and thaumatin 5 were observed to be from the increase of domain sizes within the crystals. 39 The flight samples reported in this study were consistently larger than the ground samples, and it would appear that this increase in volume is also reflected in domain sizes.…”

Protein crystal perfection and its application

“…It has been used to characterize successfully the improvement seen in some microgravity samples i.e. a reduction in the reflection mosaic spread providing a corresponding increase in the signal-to-noise ratio of the reflection (Snell et al, 1995;Helliwell et al, 1996;Ng et al, 1997). Mosaicity is the recorded rocking width of the reflection with the instrument contribution deconvoluted out.…”

Effects of Protein Purity and Precipitant Stereochemistry on the Crystallization of Thaumatin

“…29,30,44 Therefore, we crystallized the protein in 0.5 M of sodium DL-tartrate (to prepare the racemic precipitant, we mixed 0.5 M sodium L-tartrate with an equal volume of 0.5 M sodium D-tartrate). In the experiments with the racemate, we see bipyramids, prisms, and amorphous precipitate ( Figure 6); the variability in the structures that form may reflect the heterogeneous nature of the racemic solution.…”

“…The extinction coefficient of thaumatin at 280 nm was taken to be E 0.1% ) 1.27 mg mL -1 cm -1 . 28,44 Conductivity and pH measurements were performed using an Orion 4-Star conductivity and pH meter with a DuraProbe conductivity cell and a RossSure-Flow pH electrode (Thermo Fisher Scientific, Waltham, MA).…”