Shift of molecular mass after phosphorylation - How many kDa larger? (Aug/25/2006 )
We have identified a protein by MS/MS running in our gels about 56 kDa (SDS-PAGE 1D & 2D). The mass of protein without posttranslational modifications is 50 kDa, predicted from primary sequence. Its strongly phosphorylated in our system but would this result in a shift of about 6 kDa??? (of course, estimation of molecular mass in SDS-PAGE is not that reliable...); as it is a cytoplasm protein, glycosylation may not contribute to mass. Any ideas are very welcome!
what about ubiquitin? isn't that modification about 6kda.
thanks, very good idea - I just read today that it is unknown if our protein can be ubiquínilated - I will check it
i think it is possible. there is not only phosphorylation, but bunch of others modifications as well. i work with the protein which predicted mass is about 8kDa lower then the observe.. im not sure about ubiquitin.. isnt that supposed to be transient? i mean, are there proteins which are always coupled to one ubiquitin?
Sometimes phosphorylation doesn't change the way a protein runs on a gel, sometimes it can - usually only about 2kDa but if you're using a mini-gel it is difficult to discern between 2 and 6kDa (also depending on acrylamide %)