Spectroscopic characterization of Coomassie blue and its binding to amyloid fibrils
Artikel i vetenskaplig tidskrift, 2012
Coomassie brilliant blue G-250 (CB) is the dye used frequently in the Bradford assay for protein
concentration determination. In this study, we investigated how the solvent polarity and viscosity affect
the CB absorption and fluorescence spectra and apply this understanding to investigate the binding of CB
to lysozyme and insulin in the native and amyloid fibril states. Coomassie blue binds both to the native
protein and to amyloid fibrils but gives distinctly different spectral responses. The absorption and
fluorescence spectra of CB indicate that binding sites in the fibrils are less polar and hold the CB dye more
rigidly than in the native forms. The spectral comparison of CB bound to the two different fibrils showed
that the binding sites are different, and this was most likely due to differences in secondary structure as
monitored by circular dichroism. Finally, linear dichroism was used to show that the fibril-bound CB is
oriented preferentially parallel to the insulin amyloid fibril axis.
Coomassie blue
Polarity
Bradford assay
Amyloid fibrils
Viscosity
Solvent