Lysine residues in the N-terminal huntingtin amphipathic α-helix play a key role in peptide aggregation.

Abstract

Huntington's disease is a genetic neurodegenerative disorder caused by an expansion in a polyglutamine domain near the N-terminus of the huntingtin (htt) protein that results in the formation of protein aggregates. Here, htt aggregate structure has been examined using hydrogen-deuterium exchange techniques coupled with tandem mass spectrometry. The focus of the study is on the 17-residue N-terminal flanking region of the peptide that has been shown to alter htt aggregation kinetics and morphology. A top-down sequencing strategy employing electron transfer dissociation is utilized to determine the location of accessible and protected hydrogens. In these experiments, peptides aggregate in a deuterium-rich solvent at neutral pH and are subsequently subjected to deuterium-hydrogen back-exchange followed by rapid quenching, disaggregation, and tandem mass spectrometry analysis. Electrospray ionization of the peptide solution produces the [M + 5H](5+) to [M + 10H](10+) charge states and reveals the presence of multiple peptide sequences differing by single glutamine residues. The [M + 7H](7+) to [M + 9](9+) charge states corresponding to the full peptide are used in the electron transfer dissociation analyses. Evidence for protected residues is observed in the 17-residue N-terminal tract and specifically points to lysine residues as potentially playing a significant role in htt aggregation.