Aggregation of partially unfolded Myosin subfragment-1 into spherical oligomers with amyloid-like dye-binding properties.
Proteolytic myosin subfragment 1 (S1) is known to be partially unfolded in its 50-kDa subdomain by mild heat treatment at 35 degrees C [Burke et al. (1987) Biochemistry 26, 1492-1496]. Here, we report that this partial unfolding is accompanied by aggregation of S1 protein. Characteristics of the aggregate thus formed were: (i) formation of transparent sediment under centrifugation at 183,000 x g; (ii) amyloid-like, dye-binding properties such as Congo red-binding and Thioflavin T fluorescence enhancement; (iii) a uniformly sized spherical appearance in electron micrographs; and (iv) sensitivity to tryptic digestion. Gel filtration analysis of the aggregation process indicates that the spheroid was formed through an intermediate oligomeric stage. The aggregate inhibited spontaneous aggregation of an isolated 50 kDa fragment into a large amorphous mass. The remaining native regions in the partially unfolded S1 were probably responsible for this effect. These results show that, unlike the 50-kDa fragment, the partially unfolded S1 molecules do not form amorphous aggregates but assemble into spherical particles. The native regions in partially unfolded S1 may be a determinant of aggregate morphology.
Komatsu
Hideyuki
H
Department of Bioscience and Bioinformatics, Kyushu Institute of Technology, Iizuka, Fukuoka, 820-8502. hide@bio.kyutech.ac.jp
Shinotani
Nami
N
Kimori
Yoshitaka
Y
Tokuoka
Jun-Ichiro
J
Kaseda
Kuniyoshi
K
Nakagawa
Hiroyuki
H
Kodama
Takao
T
D000682:Amyloid / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D002850:Chromatography, Gel / Q000379:methods; D002942:Circular Dichroism / Q000379:methods; D003224:Congo Red / Q000737:chemistry; D006358:Hot Temperature; D007700:Kinetics; D008855:Microscopy, Electron, Scanning / Q000379:methods; D015879:Myosin Subfragments / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism; D010316:Particle Size; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011817:Rabbits; D013816:Thermodynamics; D013997:Time Factors
D000682:Amyloid; D015879:Myosin Subfragments; D009842:Oligopeptides; D010446:Peptide Fragments; D003224:Congo Red