Hi, I am working with an unknown protein. The calculated size should
be about 70 kDa. I produce it in SaOS-Cells and it is secreted into
the culture medium. It has a his and a myc tag. When I analyse it on
SDS-Page and Western Blot I always get a big and clear band at about
100 kDa and only a very thin band at 70 kDa. I know that inside the
cell it has the right size but after the secretion it is 30 kDa
larger. I already had it in MS (tryptic digestion) and only my protein
and no other protein came up. So what can it be that is bound to my
protein? We already discussed glycosylation. But people told me for
that the band is to clear. We checked with a glycosylation inhibitor
and the protein is only about 5 kDa smaller. If you have any idea what
I could check for, that would be great. Thank you very much.
