will freeze thaw cycles or sonication disrupt protein interaction? - (Feb/09/2007 )

after lyse the cells, put the lysate through freeze thaw cycles (liquid nitrogen and 37 water bath) will improve protein recovery,
But will this disrupt protein protein interactions?
How about sonication? Will it disrupt protein protein interactions?
Thanks!

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To answer the title fo the posting, I would suggest that sonicationo is more likely to disrupt protein protein interactions, because the process generates a fair bit of heat, which is never a nice thing to do to your proteins.

To clarify your posting a bit, how do you lyse the cells? Freeze-thaw shold be able to do that by itself (as will sonication, of course). I don't understand why you'd want to freeze-thaw OR sonicate *after* lysis.

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Just want to get better recovery of the potein.

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Sorry, I should have been a bit more specific with the questions. How do you lyse the cells, and what level of expression do you get in the soluble and insoluble fractions? If your expression is poor, there are a number of possible reasons from poor codon usage to toxicity of the protein you're expressing. If the total amt of protein is OK, but it's all in the insoluble fraction, then you might need to look at the expression temperature, the codon usage (again), or the way you disrupt the cells.
If you could post an image of your expression gel, showing pre- and post-induction, as well as soluble and insoluble fractions, someone here might be able to help you.
My gut instinct is to go with 2 or 3 cycles of freeze-thaw, including some lysozyme and DNase I in the lysis buffer. The technique is very gentle and still fairly quick. If you sonicate too hard or too long, you can start aggregating proteins, and possibly dissociating some complexes because of excess heating (but every protein complex could act differently....)

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