| Abstract: | In recent years, native mass spectrometry (nMS) has continued to grow in popularity and recognition as an excellent structural biology tool. The implementation of orthogonal methods to nMS, such as liquid chromatography (LC) and ion mobility spectrometry (IMS), has greatly increased the reach that MS possesses for elucidating structures of biological molecules. However, many aspects that are of vital to understanding solution-phase measurements often go overlooked when conducting nMS experiments, such as solution temperature and thermodynamic trends. Even the mechanisms of electrospray ionization (ESI) are not completely understood in respect to co-solutes and osmolytes. Herein, a newly designed, variable temperature-ESI (vT-ESI) device for modulating the solution temperature in nMS experiments is used to elucidate thermodynamic quantities for temperature-dependent protein/protein and protein/ligand interactions. The importance of osmolyte contributions toward stability of biomolecules is also underscored as these small molecules perform vitally necessary roles within the complex, dynamic cellular environment. Lastly, the mechanism of the charging of protein ions via ESI is examined in respect to how solution conditions and concentrations of analytes modulate charged ion populations. In this dissertation, methods for the examination of thermodynamic mechanisms of proteins and protein complexes will be proposed. An explanation of observed thermodynamic trends for ligand-binding proteins systems will be examined in respect to various complimentary methods. Thermodynamic explanation for the interaction mechanism(s) of osmolytes will be given which confirms aspects of osmolyte-water network interactions. Finally, evidence for a dual mechanism of ESI for charge reduction reactions will be examined in an attempt to explain how the solution environment in a charged droplet leads to drastically different charge states for protein ions. The electronic version of this dissertation is accessible from https://hdl.handle.net/1969.1/198815 |
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