Abstract
Generating high-quality crystals remains a bottleneck in biological and materials sciences. Here a counter-diffusion method was used to improve the X-ray diffraction quality of the N-terminal domain of Drosophila melanogaster Toll receptor crystals. It was observed that crystallization occurred with a peculiar pattern along the capillary resembling Liesegang bands; this phenomenon is described at both macroscopic and atomic levels. It was found that bands appeared for native protein as well as for co-crystals of magic triangle (I3C)-bound protein even though they crystallize in different space groups. Crystallization occurred with a linear recurrence independent of the precipitant concentration and a protein-specific spacing coefficient. Bandwidth varied along the capillary, oscillating between large precipitation areas and single crystals. The reported data suggest that repetitive patterns can be generated with biological macromolecules in the presence of sodium malonate as a crystallization agent. A comparison with typical Liesegang patterns and the possible mechanism underlying this phenomenon are discussed.
Highlights
Repetitive patterns such as the stripes of a zebra or those of agate rocks – very common in nature – are generated by complex mechanisms that are still only partially understood and remain difficult to model mathematically
In this study we used the N-terminal domain of Toll encompassing the 201 first amino acids produced as a variable lymphocyte receptor (VLR) hybrid protein, a strategy that can generate stable deletion constructs established for mammalian Toll-like receptors (TLRs) (Jin et al, 2007; Kang et al, 2009; Kim et al, 2007; Yoon et al, 2012)
The additional handling often resulted in crystal damage
Summary
Repetitive patterns such as the stripes of a zebra or those of agate rocks – very common in nature – are generated by complex mechanisms that are still only partially understood and remain difficult to model mathematically. Wilhelm Ostwald was the first one to propose an explanation to the phenomenon (Ostwald, 1897, 1925) His hypothesis is based on the propagation of a supersaturation wave and belongs to the pre-nucleation theories, in contrast to more recent post-nucleation theories, in which repetitive patterns are produced in essentially homogeneous and continuous colloid. Following the law of mass action, precipitation at a given position leads to a local decrease in supersaturation. An experimental setup similar to Liesegang’s experiments in gel has been used for almost two decades for protein crystallization by the method of counter-diffusion in capillaries (Garcıa-Ruiz et al, 2001; Ng et al, 2003; Otalora et al, 2009), repetitive patterns have not been observed for proteins before. We present the method used to generate material suitable for such a study and note the unusual behaviour of Toll crystals obtained by counter-diffusion crystallization in capillaries in the presence of malonate
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