Protein Crystallization Hits

Handling and storage of protein crystallization trials is usually carried out in a manner that minimizes exposure to vibration. I have set up crystallization trials and put them aside to a 'safe place' with the intention to let the crystallization commence without me interrupting the crystallization process. While I'm not sure if it was the lack of shaking or the temperature increase during observation caused by the microscope light, I did get larger and better diffracting crystals in at least one case. On the other hand, I've seen crystallization cabinets with hundreds of trays that got a whack every time somebody did not catch the door into the crystallization room. And this was a very productive protein structure lab.

Let's use this vibration effect and take it to the extreme: how about adding a bit more stress, such as ultrasonication? 3 seconds  of of bath-ultrasonication every 3 minutes throughout the crystallization process?

That's exactly what Crespo et al  have done and describe in their new paper:

Crespo, R., Martins, P., Gales, L., Rocha, F., & Damas, A. (2010). Potential use of ultrasound to promote protein crystallization Journal of Applied Crystallography, 43 (6) DOI: 10.1107/S0021889810040951

This is the contraption they used to compare with ultrasonication vs. without. 

Figure: Simple setup to compare the effect of micro batch-type protein crystallization with ultrasonication and without ultrasonication.

The setup is rather well controlled and was used to map the supersaturation curves for Lysozyme (I know…) in the presence and absence of the ultrasonication stress regime. The finding is that ultrasonication promotes nucleation. In doing so, it appears that crystals gown with this method are of higher quality. While the statistics on crystal quality (n=13) are not that convincing and the effect is small (0.1A average improvement), I'd do it if I'm desperate. For instance in cases where protein crystallization optimization by

• reagent screening, including additives,
• drop mixing regime
• temperature
• different formats and plates
• in-situ proteolysis

have not yielded any improvements in the protein crystal diffraction quality.

Cheers,

Peter