Understanding Intrinsic Biomolecular Structure with Mass Spectrometry Techniques

Over the past twenty years, since the development of “soft” ionization techniques, there has been a surge of interest in mass spectrometry based protein analysis. It is tempting to make use of mass spectrometry experiments, which are carried out in the gas phase, to elucidate biological behavior. However, the question of whether or not native structure is retained following ionization and transfer to the gas phase has yet to be definitively answered. While mass spectrometrists have made use of a variety of techniques, including fragmentation, spectroscopy, ion mobility, and ion-molecule reactions, to examine the conformation, stability, and energetics of desolvated proteins and complexes, our current understanding of gaseous biomolecules is far behind that of biomolecules in the condensed phases. This dissertation addresses the need for a gas-phase tool which is capable of providing localized conformational detail, and explores the possibility of hydrogen-deuterium (H/D) exchange as a suitable solution. Chapters 2 and 3 show data derived from H/D exchange with subsequent electron capture dissociation, which suggest that key elements of secondary structure, the β-hairpin and the α-helix, may survive electrospray. Chapter 3 also provides evidence that this combination of techniques can identify regions on a peptide that have increased conformational flexibility. In Chapter 4, a study aimed at addressing the relationship between global conformation and deuterium labelling in the gas phase is presented. The work in this thesis builds upon the existing body of gas-phase H/D exchange literature, highlighting strengths of the technique and addressing some of the challenges that users will be faced with regarding the interpretation of these data.Ph.D.2018-12-19 00:00:0