UCLA

The interrogation of protein structure, especially identifying and localizing post-translational modifications (PTMs) and binding sites of protein and ligand/small molecule interactions, is crucial for understanding protein function. Native- and native top-down mass spectrometry (nTD-MS) have become prominent analytical tools for protein characterization. The work here has addressed several experimental issues to increase the applicability of nTD-MS for elucidating protein structures. We have selected supercharging agents based on their pKa values, and demonstrated that the addition of supercharging agents can help deposit more charges on the protein ions during the ESI process. Increasing the precursor charge state enhances the efficiency of tandem mass spectrometry processes used to fragment large biomolecules. Additionally, we have investigated methods to generate sequence-specific fragments directly from protein complexes using activation/dissociation techniques alternative to traditionally used collision-based strategies, e.g., electron capture dissociation (ECD). Lastly, we applied our nTD-MS methods to aid in the determination of the higher order structure of a potential human E3 ligase. My dissertation research can contribute to advancing mass spectrometry workflows to achieve more efficient structure characterization of protein complexes, and provide complementary information for conventional structure analysis (e.g., electron microscopy, crystallography).