Effect of Trehalose on W7FW14F Apomyoglobin and Insulin Fibrillization:  New Insight into Inhibition Activity

Abstract

Trehalose, a disaccharide present in many nonmammalian species, protects cells against various environmental stresses. Trehalose has recently been shown to decrease aggregate formation and toxicity in cell models and to alleviate amyloid-induced diseases. The aim of our study was to use two amyloid-forming proteins, i.e., W7FW14F apomyoglobin and insulin, as model systems to elucidate the molecular mechanism by which trehalose affects the amyloid aggregation process and to investigate further its therapeutic potential. Protein aggregation was examined by far-UV circular dichroism, UV absorption, thioflavin T fluorescence, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, atomic force microscopy, and Fourier transform infrared spectroscopy. Cell viability was investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay. We found that trehalose does not inhibit protein aggregation but acts at different stages of the fibrillization process depending on the protein model used. In fact, trehalose dose-dependently inhibited fibril formation in the W7FW14F apomyoglobin model and increased the lag phase in the insulin model. In both cases, trehalose caused accumulation of toxic oligomeric species. The results suggest that trehalose may favor or inhibit the formation of "on-pathway" or "off-pathway" oligomeric intermediates depending on the nature of the aggregating protein.

Protein-Protein interactions leading to aggregation: Perspectives on mechanism, significance and control

“…In the same study, the effect of ectoines was shown on prevention of Aȕ 1-42 -induced cell toxicity, with ectoine being more effective [214]. On the other hand, a recent study aimed at elucidating mechanism of amyloid inhibition by trehalose involving two model proteins indicated while delay/inhibition of fibrillation are obvious, more toxic oligomeric species may be formed during the process [215]. This again highlighted the need for more detailed studies on small molecules that inhibit amyloid fibril formation.…”

Proteins in Saccharides Matrices and the Trehalose Peculiarity: Biochemical and Biophysical Properties

“…The effects is dose-dependent and depends on the protein. Also the lag phase of insuline was increased in the presence of trehalose [207].…”

External Factors Affecting Protein Aggregation

“…Additional examples include aggregation of rhGCSF at 37 ° C at pH 7 158 and W7FW14F apomyoglobin at pH 7.0 at room temperature. 120 Other proteins are prone to aggregation at a pH slightly below or above the neutral condition, including fi refl y luciferase at pH 5.5 at 30 ° C, 80 a botulinum neurotoxin (50 kD) at pH 8 at 50 ° C, 159 an Fc fusion protein at pH 3.6 at room temperature, 160 a mAb and ovalbumin at pH 4.0 at 37 ° C, 40 and a mouse mAb (mIgG1) at pH 3.9 at or above room temperature. 161 Protein aggregation caused by pH changes can be simply an effect of altered charge -charge interactions and/or pH -induced protein destabilization/ partial unfolding.…”

“…183 These sugars/polyols do not seem to work on all stages of the aggregation process. Vilasi et al 120 found that trehalose (50 -600 mM) did not suppress the early phase of the aggregation process (formation of prefi brillar oligomeric structures) of a apomyoglobin analogue but did inhibit the formation of fi brils from early aggregates in a dose -dependent manner. Because of this effect, tre- halose seems to maintain the aggregates in a nonfi brillar, highly cytotoxic state.…”