Lee, NO; (2017) Ion Mobility Mass Spectrometry Studies of Alpha1-antitrypsin. Masters thesis , UCL (University College London).

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Abstract

Mass Spectrometry (MS) has significantly contributed to the field of structural biology in the last decades. In this thesis, native MS and ion mobility-MS (IM-MS) is used to study alpha1- antitrypsin (a1-AT) in order to better understand its pathophysiology, especially in terms of aggregation, which can cause early onset emphysema and liver cirrhosis in patients with certain mutations of this protein. Initial work involved the study of recombinant a1-AT by comparing the collision cross sections (CCS) and unfolding patterns of different recombinant mutations. Two stabilising variants (T114F and G117F) and a slowly polymerising variant (K154N) were studied alongside the recombinant protein. Indeed, the proteins did show differences in their CCSs and unfolding patterns as the stabilising variants retained their native structure more than the K154N variant at high collision energies. Plasma a1-AT variants (B, Z, and S) were then studied using native and IM-MS. Native MS was able to identify certain glycan residues attached to the plasma protein whilst IM-MS confirmed the stabilising effect of glycosylation on the protein upon increases in internal energy. IM-MS also revealed that the disease variants had larger CCS values than the plasma control, which suggests that in disease a1-AT is more unfolded and primed to entering the aggregation pathway. Finally, a1-AT polymers were analysed by native MS and IM-MS. Although further work is needed, the spectra show differences in intensities of certain oligomeric species, which may lead to determining the pathway towards a1-AT aggregation.

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