Distinguishing crystal-like amyloid fibrils and glass-like amorphous aggregates from their kinetics of formation.
Amyloid fibrils and amorphous aggregates are two types of aberrant aggregates associated with protein misfolding diseases. Although they differ in morphology, the two forms are often treated indiscriminately. β(2)-microglobulin (β2m), a protein responsible for dialysis-related amyloidosis, forms amyloid fibrils or amorphous aggregates depending on the NaCl concentration at pH 2.5. We compared the kinetics of their formation, which was monitored by measuring thioflavin T fluorescence, light scattering, and 8-anilino-1-naphthalenesulfonate fluorescence. Thioflavin T fluorescence specifically monitors amyloid fibrillation, whereas light scattering and 8-anilino-1-naphthalenesulfonate fluorescence monitor both amyloid fibrillation and amorphous aggregation. The amyloid fibrils formed via a nucleation-dependent mechanism in a supersaturated solution, analogous to crystallization. The lag phase of fibrillation was reduced upon agitation with stirring or ultrasonic irradiation, and disappeared by seeding with preformed fibrils. In contrast, the glass-like amorphous aggregates formed rapidly without a lag phase. Neither agitation nor seeding accelerated the amorphous aggregation. Thus, by monitoring the kinetics, we can distinguish between crystal-like amyloid fibrils and glass-like amorphous aggregates. Solubility and supersaturation will be key factors for further understanding the aberrant aggregation of proteins.
Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
Yuichi
Y
Yoshimura
Yuxi
Y
Lin
Hisashi
H
Yagi
Young-Ho
YH
Lee
Hiroki
H
Kitayama
Kazumasa
K
Sakurai
Masatomo
M
So
Hirotsugu
H
Ogi
Hironobu
H
Naiki
Yuji
Y
Goto
D000682:Amyloid / Q000096:biosynthesis* / Q000737:chemistry*; D000817:Anilino Naphthalenesulfonates; D052160:Benzothiazoles; D004926:Escherichia coli; D005453:Fluorescence; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008854:Microscopy, Electron; D011487:Protein Conformation; D017510:Protein Folding; D057165:Proteostasis Deficiencies / Q000378:metabolism / Q000473:pathology*; D012965:Sodium Chloride / Q000737:chemistry; D013844:Thiazoles; D014465:Ultrasonics; D001613:beta 2-Microglobulin / Q000737:chemistry*
C515594:8-anilino-1-naphthalenesulfonic acid; D000682:Amyloid; D000817:Anilino Naphthalenesulfonates; D052160:Benzothiazoles; D013844:Thiazoles; D001613:beta 2-Microglobulin; C009462:thioflavin T; D012965:Sodium Chloride