How to express toxic protein in BL21(DE3) or (pLysE) strain - (Aug/06/2007 )

Hello everybody,

I'm posting this for my friend.
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If there is anybody had an experience about overexpression cytotoxic protein?

I need to overexpress a 42kd toxic protein. It did not work well in BL21(DE) strain.
I replaced the strain to BL(pLysE). I got significant amount of protein at the first time
then found the 35kd band had also been overexpressed. However, the 42kd band
disappear while the 35kd band presented.

I am wondering if there is someone who is familiar with the toxic protein overespression
could help me with this? I would really really appreciate it!!

Thank you very much

assiduoushy

You have a few options. One of the simplest is to grow the cells to a fairly high OD (up to about 1) then induce quickly, for say 1 hr, and harvest as normal.
Next option, if your protein is really toxic is to try BL21 (pLysS), because it has much tighter control of expression.
Finally, you might have to consider an intein expression system. Basically, you express the protein in two parts, then allow the two parts to self-ligate. This is really useful if you need lots of protein.
There are probably plenty of other options, but these are the first ones that came to mind.

On another matter, does your friend have any idea what the 35kDa protein band is?