The protein is 60-residue long and possesses 4 disulphide bonds.
Dear Peter,
I am not sure about cell free as I am trying it myself here with little success. It's kinda tricky I guess but again I'm new at it. However, for bacterial exp of high disulphide you can try to supplement chaperones to improve the yield of disulphide rich proteins. There are several papers on this in pubmed.
Hope this helps.. Good luck.
Dear Theam,
If possible, could you please provide reference for papers on chaperones you mentioned.
Thank you!
Dear Peter,
i do not the articles with me but if you have access maybe you can search for this two articles as a start:
PMID: 21125388
PMID: 16720693
The group does a lot of work on the chaperones.
There are ways to tweak both systems so if you haven't set up the cell-free system yet you might want to try optimisation of the E.coli system first.
You could, for example, use a strain that expresses DsbC proteins which shuffle around disulfide bonds (like the Shuffle strain from NEB: https://www.neb.com/products/c3025-shuffle-competent-e-coli).
The optmisation for disulfide bonded proteins in the cell-free system is based on the same principle in that groups have added DsbC proteins (or you could co-express) and using glutathione buffer.
read for example : http://www.ncbi.nlm.nih.gov/pubmed/23291178
Good luck!
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