Investigating Peptide-Lipid Interactions in Nanodiscs Using Native Mass Spectrometry and Fast Photochemical Oxidation of Peptides
AuthorReid, Deseree JoAnne
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PublisherThe University of Arizona.
RightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction, presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
EmbargoRelease after 10/12/2023
AbstractAntimicrobial peptides (AMPs) are an important part of the innate immune system and demonstrate promising applications in the fight against antibiotic resistant infections due to their unique mechanism of targeting bacterial membranes. Nevertheless, it is challenging to study the interactions of these peptides within lipid bilayers, making it difficult to understand their mechanisms of toxicity and selectivity. Lipoprotein nanodiscs are ideally suited for the study of AMPs with native mass spectrometry because they provide a relatively monodisperse nanoscale lipid bilayer environment for delivering membrane peptides into the gas phase. However, native mass spectrometry of nanodiscs produces complex spectra that can be challenging to assign unambiguously. This dissertation first describes a method to simplify interpretation of nanodisc spectra using an engineered series of mutant membrane scaffold proteins (MSP) that do not affect nanodisc formation but shift the masses of nanodiscs in a controllable way, thus eliminating isobaric interference from the lipids. Moreover, by mixing two different belts before assembly, the stoichiometry of MSP is encoded in the peak shape, which allows the stoichiometry to be assigned unambiguously from a single spectrum. Next, this dissertation describes the use of fast photochemical oxidation of peptides (FPOP), an irreversible footprinting technique that labels solvent accessible residues, in combination with LC-MS-MS and native charge detection-mass spectrometry to study AMP-lipid interactions and surface exposed residues within different lipid bilayer nanodiscs. The findings herein provide complementary information on the potential modes of action and lipid selectivity of various AMPs.
Degree ProgramGraduate College