Reconsidering the mechanism of polyglutamine peptide aggregation.
There are at least nine neurodegenerative diseases associated with proteins that contain an unusually expanded polyglutamine domain, the best known of which is Huntington's disease. In all of these diseases, the mutant protein aggregates into neuronal inclusions; it is generally, although not universally, believed that protein aggregation is an underlying cause of the observed neuronal degeneration. In an effort to examine the role of polyglutamine in facilitating protein aggregation, investigators have used synthetic polyglutamine peptides as model systems. Analysis of kinetic data led to the conclusions that aggregation follows a simple nucleation-elongation mechanism characterized by a significant lag time, during which the peptide is monomeric, and that the nucleus is a monomer in a thermodynamically unfavorable conformation [Chen, S. M., et al. (2002) Proc. Natl. Acad. Sci. U.S.A. 99, 11884-11889]. We re-examined this hypothesis by measuring the aggregation kinetics of the polyglutamine peptide K2Q23K2, using sedimentation, static and dynamic light scattering, and size exclusion chromatography. Our data show that during the lag time in sedimentation kinetics, there is substantial organization of the peptide into soluble linear aggregates. These aggregates have no regular secondary structure as measured by circular dichroism but have particle dimensions and morphologies similar to those of mature insoluble aggregates. The soluble aggregates constitute approximately 30% of the total peptide mass, form rapidly, and continue to grow over a period of hours to days, eventually precipitating. Once insoluble aggregates form, loss of monomer from the solution phase continues. Our data support an assembly mechanism for polyglutamine peptide more complex than that previously proposed.
Lee
Christine C
CC
Department of Chemical and Biological Engineering, University of Wisconsin, Madison, Wisconsin 53706, USA.
Walters
Robert H
RH
Murphy
Regina M
RM
D011232:Chemical Precipitation; D006816:Huntington Disease / Q000473:pathology; D008954:Models, Biological; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism*; D011108:Polymers / Q000378:metabolism*; D011485:Protein Binding; D013816:Thermodynamics
D010455:Peptides; D011108:Polymers; C097188:polyglutamine