Carbohydrate-Protein Interactions Studied Using Electrospray Ionization Mass Spectrometry
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Abstract
This thesis describes the studying of electrospray ionization (ESI) process and the application of the direct ESI-MS assay to study noncovalent carbohydrate-protein interactions in vitro. Protein unfolding induced by ESI process was first investigated. It is proposed that the Coulombic repulsion between the negatively charged residues and liquid/droplet surface charge under certain ESI conditions can induce unfolding of acidic proteins. A deleterious non-uniform response factors phenomenon induced by high molecular weight molecules and complexes in the application of direct ESI-MS assay was also investigated. It is possibly due to the reduction in the number of available surface sites in the ESI droplets upon introduction of large solute and increased competition between protein and the more hydrophilic carbohydrate-protein complex for these sites. Direct ESI-MS assay was also utilized to investigate the stepwise binding of the GM1 pentasaccharide (GM1os) to the cholera toxin B subunit homopentamer and to elucidate positive binding cooperativity.