Hi guys,
Some of my recent protein crystals have turned into sphericules. These were small crystals to begin with but they have appeared to turn into oil form. Is this common, and is there anything I can do to prevent it? My crystals are grown at 4 degrees.
Thanks again
Harry
Hi Harry, one possible reason may be that your protein is undergoing some modification, e.g. proteolysis. Did you test the stability of your protein? Otherwise it could also indicate that the conditions are still changing i.e. equilibration has not yet been reached in your experiment and due to changing solubility the crystals slowly dissolve.
To bypass the problems you may try use the initial small crystals for seeding.
Good luck, Tjaard
Hi Tjaard,
Thank you for your answer. I think seeding is something to try next. The only conditions which produced a hit included the protein incubated with its substrates. Previous SAXS studies concluded that the substrates caused a massive contraction in the protein and reduction of flexibility. So maybe the substrates are being degraded as well, resulting in a change of solubility. I know that our protein is generally less stable and only lasts a day or two once purified (without glycerol).
Thanks for your help
Ps Groningen is an excellent university. I had the opportunity to visit it the last time I was in the Netherlands and found everyone very friendly.
Hi harry, it could also be that you are using impure regants. Try crystallizing some easy protein (e.g lysozyme) with the same reagents and see what happens.
you are having liquid-liquid phase transition! check out this reference "Liquid–liquid phase separation in supersaturated lysozyme solutions and associated precipitate formation/crystallization" where the authors built an empirical phase diagram for L-L phase transition and crystallization of the same protein under virtually same solution conditions. I hope this helps
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