Spectroscopic characterization of Coomassie blue and its binding to amyloid fibrils

Nils Carlsson (Institutionen för kemi- och bioteknik, Fysikalisk kemi); Catherine Kitts (Institutionen för kemi- och bioteknik, Fysikalisk kemi); Björn Åkerman (Institutionen för kemi- och bioteknik, Fysikalisk kemi)

Analytical Biochemistry, 420 ( 1 ) s. 33-40

0003-2697

Artikel, refereegranskad vetenskaplig

2012

engelska

http://dx.doi.org/10.1016/j.ab.2011.08.043

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.

NATURVETENSKAP ->
Kemi ->
Analytisk kemi ->
Spektroskopi
NATURVETENSKAP ->
Kemi ->
Fysikalisk kemi ->
Biofysikalisk kemi
NATURVETENSKAP ->
Kemi ->
Fysikalisk kemi ->
Spektroskopi

Coomassie blue, Bradford assay, Amyloid fibrils, Solvent, Polarity, Viscosity

Livsvetenskaper

Grundläggande vetenskaper

147479

146896

2011-10-20 11:40