Simultaneous monitoring of peptide aggregate distributions, structure, and kinetics using amide hydrogen exchange: application to Abeta(1-40) fibrillogenesis.
Increasing evidence indicates that soluble aggregates of amyloid beta protein (Abeta) are neurotoxic. However, difficulty in isolating these unstable, dynamic species impedes studies of Abeta and other aggregating peptides and proteins. In this study, hydrogen-deuterium exchange (HX) detected by mass spectrometry (MS) was used to measure Abeta(1-40) aggregate distributions without purification or modification that might alter the aggregate structure or distribution. Different peaks in the mass spectra were assigned to monomer, low molecular weight oligomer, intermediate, and fibril based on HX labeling behavior and complementary assays. After 1 h labeling, the intermediates incorporated approximately ten more deuterons relative to fibrils, indicating a more solvent exposed structure of such intermediates. HX-MS also showed that the intermediate species dissociated much more slowly to monomer than did the very low molecular weight oligomers that were formed at very early times in Abeta aggregation. Atomic force microscopy (AFM) measurements revealed the intermediates were roughly spherical with relatively homogenous diameters of 30-50 nm. Quantitative analysis of the HX mass spectra showed that the amount of intermediate species was correlated with Abeta toxicity patterns reported in a previous study under the same conditions. This study also demonstrates the potential of the HX-MS approach to characterizing complex, multi-component oligomer distributions of aggregating peptides and proteins.
Qi
Wei
W
Department of Chemical Engineering, University of Virginia, Charlottesville, Virginia 22904, USA.
Zhang
Aming
A
Patel
Dhara
D
Lee
Sungmun
S
Harrington
Jamie L
JL
Zhao
Liming
L
Schaefer
David
D
Good
Theresa A
TA
Fernandez
Erik J
EJ
D000577:Amides / Q000737:chemistry*; D000682:Amyloid / Q000737:chemistry*; D016229:Amyloid beta-Peptides / Q000032:analysis* / Q000737:chemistry* / Q000648:ultrastructure; D041961:Deuterium Exchange Measurement; D007700:Kinetics; D013058:Mass Spectrometry; D018625:Microscopy, Atomic Force; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry* / Q000648:ultrastructure; D017433:Protein Structure, Secondary
D000577:Amides; D000682:Amyloid; D016229:Amyloid beta-Peptides; D010446:Peptide Fragments; C078592:amyloid beta-protein (1-40)