Author Type

Graduate Student

Date of Award

Summer 7-29-2026

Document Type

Thesis

Publication Status

Version of Record

Submission Date

August 2026

Department

Chemistry and Biochemistry

College Granting Degree

Charles E. Schmidt College of Science

Department Granting Degree

Chemistry and Biochemistry

Degree Name

Master of Science (MS)

Thesis/Dissertation Advisor [Chair]

Maré Cudic

Abstract

Alzheimer’s disease (AD) is characterized by the accumulation of extracellular amyloid-β (Aβ) plaques and intracellular neurofibrillary tangles composed of aggregated tau protein. Among the numerous post-translational modifications that regulate tau function, phosphorylation and O-GlcNAcylation have emerged as key modulators of tau aggregation and pathology. While phosphorylation is generally associated with tau misfolding and fibril formation, O-GlcNAcylation has been proposed to exert protective effects through its interplay with phosphorylation. However, the site-specific structural consequences of these modifications remain incompletely understood.

The objective of this work was to develop synthetic tools for the site-specific incorporation of O-GlcNAc and phosphate modifications into tau peptide models and to evaluate how these modifications influence aggregation behavior. A β-GlcNAc-serine pentafluorophenyl ester building block was synthesized and incorporated into tau-derived peptides using Fmoc solid-phase peptide synthesis. Five peptide models corresponding to residues I392–A426 of human tau were prepared, including unmodified tau and analogs modified at Ser396 or Ser400 with either O-GlcNAc or phosphate. Peptides were purified and characterized by HPLC, MALDI-TOF mass spectrometry, and NMR spectroscopy. Structural and aggregation properties were investigated by circular dichroism (CD) spectroscopy and transmission electron microscopy (TEM) in the absence and presence of Aβ40.

CD spectroscopy revealed modification- and solvent-dependent differences in the spectral profiles of the tau peptide models. All peptides retained substantial random-coil character, while the unmodified and phosphorylated variants displayed broader long-wavelength features consistent with mixed ordered contributions whereas the O-GlcNAcylated variants retained more random-coil-like spectral profiles. TEM analysis demonstrated that phosphorylation promoted the formation of extended fibrillar assemblies, particularly at Ser396, whereas O-GlcNAcylation produced more compact and less ordered aggregate morphologies. Site-specific effects were most pronounced at Ser396, indicating that this residue plays a greater role in directing aggregation behavior within the PHF-1 region. Coincubation with Aβ40 further enhanced fibrillar assembly of phosphorylated peptides, while O-GlcNAcylated peptides maintained distinct aggregate morphologies with reduced incorporation into extensive fibrillar networks.

Collectively, these findings demonstrate that phosphorylation and O-GlcNAcylation direct tau aggregation along distinct structural pathways. Rather than simply suppressing aggregation, O-GlcNAcylation appears to alter the mode of assembly, favoring alternative aggregate states that differ from the fibrillar structures promoted by phosphorylation. The synthetic methodology and peptide models developed in this work provide useful tools for investigating the molecular relationship between post-translational modification and tau aggregation in Alzheimer’s disease.

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