Record in bacterial expression of cysteine rich proteins ?

Dear Colleagues,

I am wondering how you deal with cysteine rich proteins (in the range of 10-20,) in bacterial expression ( I don' t mean small proteins that Renaud published)

In know that some labs even don't touch these bacterial constructs and go for eucaryotic directly. We frequently get cysteine rich targets, and in some cases could express soluble protein in bacteria.

But anytime I am confronted with such a project I am sceptical, especially if nothing is known about the protein, in particular S-S bonding.

I would be very interested in your epxerience !

Best

Sabine

Posted on 12-Jun-2014 16:26 CEST

Dear Sabine,

From our humble experience with bacterial expression of large cys rich proteins, we consider trying the expression in bacteria only if the protein is a cytosolic protein, with mostly reduced cysteines, and not natively secreted. forming multiple disulfide bonds in the cytosol stands very little chance of correct disulfide bonds formation... 

We had some sporadic luck with TRX fusion, but again, with not more than 4-6 disulfide bonds. 

For secreted proteins we immediately turn to insect cells, where we have much better results & faster.

Good luck,

Tsafi.

 

 

 

Posted on 12-Jun-2014 18:39 CEST

Like Tsafi, I also think that your chance to correctly express it in bacteria is extremely low. 

 

Mario

Posted on 13-Jun-2014 11:52 CEST

Like Tsafi, I also think that your chance to correctly express it in bacteria is extremely low. 

 

Mario

Posted on 13-Jun-2014 11:52 CEST
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