(230c) Examining Polyglutamine Peptide Length: A Connection Between Collapsed Conformations and Increased Aggregation

Abnormally expanded polyglutamine domains in proteins are associated with several neurodegenerative diseases, of which the best known is Huntington's. The affected proteins share no sequential, compositional, or structural homologies, with the exception of the polyglutamine domain. Expansion of the polyglutamine domain facilitates aggregation of the affected protein, and several studies directly link aggregation to neurotoxicity. The age of onset of disease is inversely correlated with the length of the polyglutamine domain; this correlation motivates an examination of the role of the length of the domain on aggregation. In this investigation, peptides containing 8 to 24 glutamines (Q8-Q24) were synthesized, and their conformational and aggregation properties were examined. All peptides lacked regular secondary structure as determined by circular dichroism. Fluorescence resonance energy transfer (FRET) studies revealed that the peptides became increasingly collapsed in phosphate buffered saline as the number of glutamine residues increased. The effective persistence length was estimated to decrease from ~11 Å to ~8 Å as the number of glutamines increased from 8 to 24. Further analysis of the data indicated that water is a good solvent for Q8 and Q12, a theta solvent for Q16, and a poor solvent for Q20 and Q24. By dynamic light scattering, we observed that Q16, Q20, and Q24, but not Q8 or Q12, immediately formed soluble aggregates upon dilution into phosphate buffered saline at 37°C. Thus, Q16 stands at the transition point from good to poor solvent, and from stable to aggregation-prone peptide, in agreement with recent theoretical simulations (Vitalis et al., J. Mol. Biol. 2008, 384:279-297). Within the set of aggregating peptides, the amount of aggregated material was found to increase with increasing length of the polyglutamine domain. Examination of aggregates by transmission electron microscopy, along with kinetic assays for sedimentation, provided evidence indicating that soluble aggregates mature into sedimentable aggregates, with maturation occurring more quickly with increasing length of the polyglutamine domain. Together, the data support a mechanism of aggregation in which monomer collapse is accompanied by formation of soluble oligomers; these soluble species lack regular secondary structure but appear morphologically similar to the sedimentable aggregates into which they eventually mature.