Hello there trypsin
it could be several things as you mentioned
1) cloning problem or addition of extra sequence into the protein (or new start site / or stop site)
2) addition of glycosylation residues on the protein (phosphorylation wouldnt increase it that big
3) ladder issues with your protein marker (maybe you mistakened it for the wrong site)
4) salt issues / buffer issues can easily change the estimated size of your protein
5) aggregation or binding to another protein? possibly due to bad lysis/lack of beta-mercaptoethanol in sample / lysis buffer.
I am out of ideas for more possibilities... anyone else?
please tell us what happened.