X23 - Self-assembly Behavior of Aggregation-prone Tau Sequences Conjugated to Amphipathic Peptides

Saturday, October 11, 2025

10:00 AM - 11:00 AM PDT

Location: Exhibit Hall F, G & H

Presenting Author(s)

LA

Leili Asgharzadeh Falbinan

Undergraduate Researcher
University of Arizona
Tucson, Arizona, United States

Co-Author(s)

Darcy R. Harris, BA

PhD Candidate
Washington University in St. Louis
Clayton, Missouri, United States
PS

Piyoosh Sharma

Research Assistant Professor
Washington University in St. Louis, United States
MJ

Meredith Jackrel

Primary Investigator
Washington University in St. Louis, United States

Last Author(s)

Jai Rudra

Associate Professor
Washington University in St. Louis, Missouri, United States

Primary Investigator(s)

Jai Rudra

Associate Professor
Washington University in St. Louis, Missouri, United States

Introduction: : Tauopathies are a group of neurodegenerative diseases, such as Alzheimer’s, caused by the aggregation of tau protein into amyloid fibrils. Within tau, two regions prone to aggregation drive this process, with the VQIINK hexapeptide being a particularly aggressive motif known to form tightly packed steric zippers made of beta sheets1. These fibrils display distinct structural “strains” that can influence disease progression and complicate diagnosis. This project aims to inhibit tau aggregation by using KFE8, a self-assembling, non-toxic peptide already explored for biomedical applications and as a vaccine scaffold2. By synthetically linking KFE8 to either end of VQIINK, we hope to develop an inhibitor that can disrupt the aggregation pathway and reduce fibril formation. This approach will help improve understanding of tau aggregation mechanisms and open new avenues for targeting tauopathies.

Materials and

Methods: : Solid phase peptide synthesis was employed to synthesize peptides Ac-VQIINK-Am, Ac-FKFEFKFE-Am, Ac-FKFEFKFESGSGVQIINK-Am, and Ac-VQIINKSGSGFKFEFKFE-Am. The peptides were cleaved, deprotected, and purified to over 90% purity using HPLC. Their mass was confirmed by MALDI-TOF mass spectrometry. Then, the peptides were dissolved in ultrapure water at 2 mM, followed by dilution to a final concentration of 0.5 mM in buffer 2 PBS with MgCl₂, 40 mM heparin, and 5 mM DTT to emulate tau aggregation conditions in vitro. Parallel samples without heparin and DTT were prepared to assess spontaneous aggregation. The 0.5 mM assay was performed with multiple replicates. For each assay, 95 μL of peptide sample was mixed with 5 μL of Thioflavin T (ThT), a fluorescent dye that binds to amyloid fibrils and exhibits increased fluorescence when bound. Samples were incubated at 37°C for five days, and ThT fluorescence was measured every 24 hours using a microplate reader to monitor fibril formation over time.

Results, Conclusions, and Discussions:: The ThT assays indicated aggregation in all samples, evidenced by increased fluorescence. The buffer negative control confirmed this increase and validated the aggregation results. Prior research shows that KFE8 and VQIINK tend to aggregate and form amyloid fibrils, making them the primary positive controls. To evaluate the impact of terminus positioning, KFE8-VQIINK and VQIINK-KFE8 were analyzed further. VQIINK-KFE8 displayed significantly higher aggregation than KFE8-VQIINK on most days, except day 0 where no difference was observed. The effect of heparin as an inducer was also tested to determine if VQIINK alone or linked to KFE8 would aggregate; no significant differences were found with or without heparin across all days. These results highlight the role of KFE8 at both N- and C-termini of VQIINK in aggregation during a 5-day ThT assay. Additional experiments, including fibril visualization with Transmission Electron Microscopy (TEM), are needed to confirm inhibition effects. Variants of KFE8 and VQIINK can be synthesized to alter fibril formation further and improve aggregation inhibition. Overall, the study concludes that peptide conjugates, especially those linking KFE8 to VQIINK, influence tau fibril formation, with effects depending on peptide configuration and terminus placement. Future research will involve additional trials, fibril visualization, and modifications to enhance strategies against tau aggregation in neurodegenerative diseases.

Acknowledgements and/or References (Optional): : 1. Seidler, P. M., Boyer, D. R., Rodriguez, J. A., Sawaya, M. R., Cascio, D., Murray, K., Gonen, T., & Eisenberg, D. S. (2018). Structure-based inhibitors of tau aggregation. Nature Chemistry, 10(2), 170–176. https://doi.org/10.1038/nchem.2889.
2. Clover, T. M., O’Neill, C. L., Appavu, R., Lokhande, G., Gaharwar, A. K., Posey, A. E., White, M. A., & Rudra, J. S. (2020). Self‑assembly of block heterochiral peptides into helical tapes. Journal of the American Chemical Society, 142(47), 19809–19813. https://doi.org/10.1021/jacs.9b09755.