Collision cross sections of protein ions and noncovalent complexes of proteins studied by electrospray ionization-mass spectrometry

The advent of gentle ionization sources has made it possible to study the higher-order structure of proteins, and noncovalent interactions between proteins, and small molecules or other proteins with mass spectrometry (MS). Electrospray ionization (EST), which allows formation of intact gas phase protein ions, is one such technique. This work focuses on fundamental aspects of the structure and stability of gas phase proteins formed by ESI.; Collision cross sections, a measure of ion “size”, provide insight into the conformations of protein ions. The first part of this thesis concerns the experimental and theoretical descriptions of collision cross sections of gas phase protein ions. The results were interpreted with a drag coefficient model, in which the thermal motion of the target gas, the scattering angle distribution, and inelastic collisions have been effectively included by introducing a drag coefficient to a previously proposed simple hard sphere model. The drag coefficient model suggests the projection areas obtained from the ion mobility should be reduced by a factor of about 0.74. Cross section results obtained from the energy loss method and the ion mobility method agree within 3% when both are interpreted with the diffuse scattering model. This work has successfully unified two methods of cross section measurements, energy loss and ion mobility.; The second part of this work develops and evaluates a new collision model which can be used to calculate relative energies transferred to protein ions in tandem mass spectrometry. This collision model considers the collision cross sections and the energy losses of ions in the activation process in the collision cell of a triple quadrupole tandem mass spectrometer system.; Noncovalent complexes of proteins binding a small molecule or another protein were studied using the approaches developed in this work. In the case of myoglobin, highly charged holomyoglobin ions were observed by ESI-MS with a novel continuous-flow mixing setup. Collision cross section measurements show that the protein has unfolded appreciably in high charge states. However, measurements of the energies needed to dissociate heme show that the heme binding energy decreases only slightly in these more highly charged ions. Further, noncovalent interactions of bovine liver cytochrome b ₅ and a series of yeast iso-1-cytochrome c mutants were studied. The results show that similar energies are required to dissociate gas phase complexes of these mutants with cytochrome b₅. (Abstract shortened by UMI.)...