Gas-Phase Conformations Of A Huntingtin N-Terminal Peptide Reveal Condensed-Phase Heterogeneity With And Without The Presence Of A Ppii Helix

Huntingtin aggregate morphology and kinetics are modulated by the presence of two flanking sequences: a seventeen-residue α-helix (Nt17) which lies N-terminal to the amyloidogenic polyglutamine region; and a polyproline PPII helix that is C-terminal to the polyglutamine region. Nt17 is responsible for aggregate nucleation, and as such, represents an intriguing target for gaining structural insight into the early stages of N-terminal huntingtin aggregation. This study examined the secondary, tertiary, and quaternary arrangement of Nt17 using ion mobility-mass spectrometry (IMS-MS) coupled with condensed-phase covalent modification and gas-phase isotopic labeling. Monomeric Nt17 adopted two gas-phase conformations, which were derived from solution structures. These structures ranged from compact globular to elongated helical. Nt17 multimers followed the same pattern, again adopting structures varying from non-specific globule to bundled helix. Species ranging from the monomer up to the pentamer were observed. Covalent modification studies reveal threonine-3 and lysine-6 are solvent-exposed in the multimeric form. Additionally, polyproline, in a PPII helix conformation, was incubated with Nt17. Gas-phase isotopic labeling studies (hydrogen-deuterium exchange, HDX) on the two non-covalent complexes revealed nearly the same amount of deuterium uptake per Nt17 monomer in the complex, which suggests the same binding face is involved in Nt17 multimer and Nt17-Polyproline interactions. These results provide structural insight into Nt17 multimerization, and thus, the early stages of N-terminal huntingtin aggregation.