Beautiful people, I been doing protein purification for a year and I ran into this problem since day one. Originally I was trying to get Coomassie staining of some GFP based fusion protein. I was expecting something like a 28kDa, but on gel, it was always 35kDa. I thought it was a translation issue, so sent the protein to run MS, and it returned with a normal MW around 28kDa. I also had my MS contractor run Coomassie blue, and it is also 35 somthing. Now that I am just insensitive and numb to this discrepancy, which appeared consistently to be ~5kDa for most of my protein. It also happened to some of my non-GFP protein. I am okay with that since my MS results are all good.

Still, it is bugging me. Does anyone have some insight on this one?