p65-HaloTag copurifying proteins in human 293T cells

In effort to develop methodology for targeted top down mass spectrometry of NF kappa B p65 from human cells, we evaluated the utility of HaloTag for purification and analysis of recombinant protein. During our study, two datasets of bottom up LC-MS/MS were generated: one from in-gel digestion of the predominant band following p65-HaloTag purification, another from in-solution digestion of all the proteins present in a p65-HaloTag purification. p65-HaloTag copurifying proteins identified in our datasets include the known interactors c-Rel, NF-kappaB p105, NF-kappaB p100, and NF-kappaB inhibitor beta. Over 100 proteins were identified by at least two peptides using a Mascot ion cut-off score of 30.