Patent ID: 8669116

Claim:
A method for determining a relative abundance of ubiquitination in a plurality of ubiquitinated polypeptides in biological samples, the method comprising: contacting cellular polypeptides of samples from each of a first cell and a second cell with a binding partner that binds to at least one molecule selected from the group consisting of: a ubiquitin molecule and a molecule linked to ubiquitin, thereby forming ubiquitinated polypeptide:binding partner complexes; isolating and separating the ubiquitinated polypeptide:binding partner complexes in the samples from the first cell and the second cell; digesting the ubiquitinated polypeptides of the complexes with trypsin, thereby generating a plurality of test peptides; determining the presence of at least one isopeptide bond between ubiquitin and an internal lysine residue of at least one of the plurality of test peptides by mass spectrometry by observing at least one mass change of 114 Daltons corresponding to a Gly-Gly residue linked to the internal lysine, wherein the presence of the bond indicates a site or sites of ubiquitination, thereby identifying the ubiquitination site or sites in the first cell and in the second cell; and, analyzing the abundance of ubiquitination in the first cell and the second cell by comparing numbers of the ubiquitination site or sites in each of the first cell and in the second cell.