I purified a mutant version of EF-Tu (E348C) that has a his-tag and an amino acid sequence that TEV protease cleaves to remove the his-tag.
I purified by Ni-NTA column and added about 40% glycerol to my samples. They are stored in a buffer containing 20 mM Tris-HCl at pH 7.5, 300 mM NaCl, 1mM beta-mercaptoethanol, and 250 mM imidazole (this is the elution buffer from the column).
I completely forgot to put them in the -20 degree celsius freezer before leaving yesterday and do not have access to the building again until tomorrow morning. It looks like they will be at 4 degrees celsius in this buffer for a total of 36 hours or so.
I know beta-mercaptoethanol is not stable in solution, but are my proteins likely to be dead after this period of time?
BME is dead for sure. Now, coming to the protein - It is not protein- RNA complex then may be you are fine. 36 Hrs is quite long and answer of your question is : Yes and No - It really depends on the protein itself, some proteins can stay for month, some few hours, some dont want to stay :)-
Better judge for this query are the people who handled this protein and know about the stability of this species.
As of now , following is the quick biochem solution for you:
What you have to do is : You have his tagged protein then simply do the WB, and check if you find a clear single band like what you saw in earlier preps
@ Mahesh Chandak,
Thanks for the reply! I had a feeling it would depend on the protein.
I also just remembered that my purification and fluorescent labeling procedures require several dialyses that are about 22 hours in length at 4 degrees, and I have never seen the protein precipitate or die during those so I am probably good.
Keeping protein for dialysis is an entirely different process, here your protein is in imidazole, exposing it to the imidazole for long time can precipitate your protein. As protein is already eluted out now you may increase EDTA concentration.
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