Ubiquitination/SUMOylation

A major mode of protein degradation involves tagging of proteins by multiple ubiquitin proteins followed by delivery of tagged proteins to the proteasome where the protein is degraded. Mono- and poly-ubiquitination is also a major post-translational modification that is involved in many cellular processes including organelle and receptor trafficking. The approach for locating sites of ubiquitination/SUMOylation is to perform a tryptic digestion of the sample(s) of interest prior to direct infusion onto the FT-ICR MS with subsequent MS/MS on detected ubiquitinylated peptides or to inject the digested sample via LC-MS/MS on an LTQ Orbitrap MS instrument followed by database search.

The process of tryptic digestion on a ubiquitinylated protein leaves behind a Gly-Gly (mass shift of +114.042927, monoisotopic) residue from ubiquitin that is bound to the Lys of the modified protein. platforms). For quantitative analysis, DIGE, Label-Free, or iTRAQ platforms can be sued prior to the digestion (Patel et al, 2009).

Patel J.C., Hueffer K., Lam T.T., Galán J.E. (2009) Diversification of a Salmonella virulence protein function by ubiquitin-dependent differential localization. Cell. 137(2):283-294.