Graduation Term

Summer 2025

Degree Name

Master of Science (MS)

Department

Department of Chemistry

Committee Chair

George L. Barnes

Committee Member

Raynard Shawn Hitchcock

Committee Member

Susil Baral

Abstract

In tandem mass spectrometry (MS/MS), chemical structure strongly influences ionization and fragmentation, yielding distinctive spectral patterns. However, standard MS/MS spectra only capture precursor and product ion’s mass to charge ratio, providing limited insight into the transient intermediates and dynamic events that govern dissociation. The effects of chemical modifications, such as post-translational modifications (PTMs) like phosphorylation, can redirect fragmentation pathways and obscure spectral interpretation.

Computational simulations can help provide insight into these processes; however, such simulations produce a vast amount of data. To address this, we developed a graph theory-based framework that enables automated, high-resolution analysis of direct dynamics simulations under collision-induced dissociation (CID). Molecular structures are encoded as augmented graphs, incorporating both connectivity and molecular property and particular to this work formal charge information. This enables the systematic identification of transient species and the construction of ensemble reaction graphs (ERGs) that comprehensively represent sampled fragmentation pathways. Filtering techniques and RDkit visualizations allow the extraction of chemically meaningful mechanisms.

This approach is applied to N-terminus protonated threonine (Thr-H⁺) and N-terminus protonated O-phosphorylated threonine (p-Thr-H⁺), revealing how phosphorylation and methylenation alters fragmentation behavior. Simulations for Thr-H⁺ reproduced key experimental peaks (m/z 102, 74, 56), with m/z 102 arising from water loss via the side-chain hydroxyl and m/z 74 and 56 resulting from loss of H2O + CO and loss of 2H2O + CO, respectively. Simulations show that m/z 74 dominates at the moderate collision energies. Phosphorylation at the side-chain hydroxyl shifted the dominant fragment to m/z 102, now corresponding to H3PO4 loss. Two isomers were identified: one via a three-membered SN2-type mechanism and another from β-elimination. Unlike phosphorylated serine, which favors β-elimination, p-Thr-H⁺ preferentially forms the three-membered ring SN2 product at moderately high collision energies due to its additional CH2 group, a trend aligning with recent experimental data. Unique to p-Thr-H⁺ is the roaming-mediated formation of m/z 99 ([H4PO4]⁺), highlighting complex charge-migration behavior. Additional fragments such as m/z 182, 154, 58, and 74 further emphasize how small structural changes and modifications open new reaction channels.

Comparisons with serine-based analogs underscore the impact of small structural differences, such as a single methylene group, on fragmentation preference and product stability. These findings demonstrate how the integration of direct dynamics with graph-based analysis provides a mechanistically insightful platform for studying peptide fragmentation, particularly for chemically modified systems where standard MS/MS interpretation is limited.

Access Type

Thesis-Open Access

DOI

https://doi.org/10.30707/ETD.1763755358.966647

Download

Available for download on Friday, September 10, 2027

Share

COinS