my contract has his-MBP-Membrane protein when i tray to express it goes in to inclusion body. for purification i lysed cell pellet with 6M GuHCl and 0.15%tritonx100 then over column i exchange tritonx100 to 0.1% of DDM and then eluted in presence of 0.1% DDM and 6 M GuHCl
for refolding of protein dialysed
1. 4M GuHCl in PBS pH7.0 for 6hr
2. 2M GuHCl in PBS pH7.0 for 4hr
2. 2M Urea in TrisHCl(50mM) pH7.0 for 6hr
protein is slightly precipitated in 4 M GuHCl and almost completely in 2M GuHCl
If you have 6xHis tag in your construct, you can try to refold your protein on the column:
https://www.gelifesciences.co.jp/catalog/pdf/18113437ac.pdf
but it can refolds the membrane protein part ?
Because in dialysis also concentration of GuHCl is decrease slowly from 6M to 4M and then from 4M to 2M GuHCl.
you tried this refolding with your protein ?
I did try it successfully with a few proteins that formed inclusion bodies, but none of them were membrane proteins.
That depends on the type of the protein. For membrane-associated proteins (type 5 and possibly type 6) your protocol has a chance to work. For integral membrane proteins (type 1, 2, 3 and 4), however, translation, folding and membrane integration occur concurrently via SRP and its receptor. They contain "annular" lipids that are mandatory part of their structure. Expressing these proteins in cytosol will not result in functional proteins.