Effect of binding of Calcofluor White on the carbohydrate residues of alpha1-acid glycoprotein (orosomucoid) on the structure and dynamics of the protein moiety. A fluorescence study.
Calcofluor White is a fluorescent probe that interacts with polysaccharides and is commonly used in clinical studies. Interaction between Calcofluor White and carbohydrate residues of alpha1-acid glycoprotein (orosomucoid) was previously studied at low and high concentrations of Calcofluor compared to that of the protein. alpha1-Acid glycoprotein contains 40% carbohydrate by weight and has up to 16 sialic acid residues. At equimolar concentrations of Calcofluor and alpha1-acid glycoprotein, the fluorophore displays free motions [Albani, J. R.; Sillen, A.; Coddeville, B.; Plancke, Y. D.; Engelborghs, Y. Carbohydr. Res. 1999, 322, 87-94], while at high concentration of Calcofluor, its surrounding microenvironment is rigid, inducing the rigidity of the fluorophore itself [Albani, J. R.; Sillen, A.; Plancke, Y. D.; Coddeville, B.; Engelborghs, Y. Carbohydr. Res. 2000, 327, 333-340]. In the present work, red-edge excitation spectra and steady-state anisotropy studies performed on Trp residues in the presence of Calcofluor, showed that the apparent dynamics of Trp residues are not modified. However, deconvoluting the emission spectra with two different methods into different components, reveals that the structure of the protein matrix has been disrupted in the presence of high Calcofluor concentrations.
Albani
J R
JR
Laboratoire de Biophysique Moléculaire, Université des Sciences et Technologies de Lille, Bâtiment C6, 59655, Villeneuve d'Ascq, France. jihad-rene.albani@univ.lille.fr
D016880:Anisotropy; D001557:Benzenesulfonates / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002241:Carbohydrates / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D009961:Orosomucoid / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013696:Temperature; D014364:Tryptophan / Q000737:chemistry*
D001557:Benzenesulfonates; D002241:Carbohydrates; D005456:Fluorescent Dyes; D009961:Orosomucoid; C007061:C.I. Fluorescent Brightening Agent 28; D014364:Tryptophan