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March 16, 2026
Feixuan Wu, Pharmsci graduate student (Li Lab), will be defending her PhD research thesis:
Advances in Mass Spectrometry-Based Proteomics and Post-translational Modification Analysis: Method Development and Biomedical Applications
Mass spectrometry (MS) has emerged as a central platform for comprehensive protein characterization and post-translational modification (PTM) analysis, benefiting from its high speed, resolution, and sensitivity. Coupling liquid chromatography with tandem mass spectrometry (LCMS/MS) enables global profiling of proteomes and PTMs, providing powerful molecular insights that advance disease mechanism studies and biomarker discovery. This dissertation contributes to MS-based proteomics through the development of innovative analytical workflows and their application to diverse biomedical models.
First, I developed a novel multiplexed strategy to enable simultaneous enrichment and quantitative analysis of N-glycopeptides and phosphopeptides, using epoxy-ATP-Ti4+-Immobilized Metal Affinity Chromatography (IMAC) in combination with N,N-dimethyl leucine (DiLeu) isobaric tags. This dual-functional platform enables coordinated interrogation of glycosylation–phosphorylation crosstalk from limited biological samples. The workflow was successfully applied to Alzheimer’s disease (AD) and diabetes models. To further expand multiplexed PTM analysis, a parallel quantitative workflow integrating global proteomics and N-glycoproteomics was established and applied to human benign prostatic hyperplasia (BPH) tissues and progeria mouse brain samples. To address the analytical challenges of O-glycosylation, a high-throughput multiprotease strategy was developed to enhance O-glycosite localization and O-glycan structural characterization in pancreatic tissues. This approach significantly improves site-specific
resolution and expands coverage of mucin-type O-glycosylation in complex biological samples. Additionally, to investigate proline hydroxylation, which is a critical modification in extracellular matrix (ECM), a quantitative workflow integrating S-Trap, optimized hydrophilic interaction chromatography (HILIC), and isobaric labeling was established. Application of this method
generated the most comprehensive site-specific hydroxyproline datasets reported to date in human pancreas, providing new insights into ECM remodeling in pancreatic disease. Collectively, this dissertation presents methodological innovations that enhance sensitivity, multiplexing capability, and PTM coverage in MS-based proteomics. By applying these advances across neurodegenerative, metabolic, and prostatic disease models, this dissertation highlights the broad utility of integrative proteomic strategies for mechanistic discovery and translational biomedical
research.