Exploring the Aggregation Propensity of PHF6 Peptide Segments of the Tau Protein using Ion Mobility Mass Spectrometry Techniques

Peptide and protein aggregation involves the formation of oligomeric species, but the complex interplay between oligomers of different conformations and sizes complicates their structural elucidation. Using ion mobility mass spectrometry (IM-MS), we aim to reveal these early steps of aggregation for the Ac-PHF6-NH2 peptide segment from tau protein, thereby distinguishing between different oligomeric species, and gaining an understanding of the aggregation pathway. An important factor that is often neglected, but which can alter the aggregation propensity of peptides, is the terminal capping groups. Here we demonstrate the use of IM-MS to probe the early stages of aggregate formation of the Ac-PHF6-NH2, Ac-PHF6, PHF6-NH2, and uncapped PHF6 peptide segments. The aggregation propensity of the four PHF6 segments is confirmed using thioflavin T fluorescence assays and transmission electron microscopy. Post-IM fragmentation and quadrupole selection on the TIMS-Qq-ToF (trapped ion mobility) spectrometer are introduced to improve oligomer assignment. In addition, TIMS collision cross section values are compared with travelling wave ion mobility (TWIMS) data to evaluate potential instrumental bias in the trapped ion mobility results. The two IM-MS instrumental platforms are based on different ion mobility principles and have different configurations, thereby providing us with valuable insight into the preservation of weakly bound biomolecular complexes such as peptide aggregates