I found many different protocols for TEV protease growth. In some of them the overexpression is overnight at low temp (18 C) and in some of them 5 hr of overexpression at 37 C. I would like to use the overnight overexpression but not sure if it can affect the yield.
I typically expressed TEV overnight at low temperatures and got very good yields. I've never done a comparison with 37C expression for 5 hrs, so I don't know which is better, but you will get a lot of usable enzyme expressing at 18C (from a 9L batch of cells, we typically got well over 100mg of enzyme).
It depends upon concentration of IPTG and post-induction temperature. You can get sufficient amount of soluble proteins by inducing with low concentration of IPTG at low temperature overnight. However, induction of TEVp expression with high concentration of IPTG at high temperature often results in formation of inclusion bodies.
Thank you for your response!
In the protocol I found with the low temperature, the IPTG concentration is 0.1mM. Is it a good concentraion? What is the recommended concentration?
0.1 mM IPTG will work good. I used 0.1 mM IPTG at 18 degree for expression of hydrophobic proteins. I got sufficient amount of soluble proteins.
I guess you were concerned about the TEV cleaving each other during longer over-expression period, resulting in useless purified TEV dead body? LOL. I am also interested in this factor and I guess none of the replies actually cared this aspect. I will keep you posted once I purified and carried out cleavage, and it will be 1-2 weeks.
I usually induce at an OD600 of 1.0, 0.5mM IPTG, 25 C for 8-12 hrs, high temp may reduce the enzyme activity, low temp reduce the productivity of TEV. Also gel filtration with 20Tris 300mM NaCl pH7.6 may improve TEV purity. For storage, I usually supply glycerol to 10%, and store the TEV in -80C.
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