Protein-flavonol interaction: fluorescence spectroscopic study.
Recent studies have shown that various synthetic as well as therapeutically active naturally occurring flavonols possess novel luminescence properties that can potentially serve as highly sensitive monitors of their microenvironments in biologically relevant systems. We report a study on the interactions of bovine serum albumin (BSA) with the model flavonol 3-hydroxyflavone (3HF), using the excited-state proton-transfer (ESPT) luminescence of 3HF as a probe. Upon addition of BSA to the flavonoid solutions, we observe remarkable changes in the absorption, ESPT fluorescence emission and excitation profiles as well as anisotropy (r) values. Complexation of 3HF with protein results in a pronounced shift (20 nm) of the ESPT emission maximum of the probe (from lambda(max)(em) = 513 nm to lambda(max)(em) = 533 nm) accompanied by a significant increase in fluorescence intensity. The spectral data also suggest that, in addition to ESPT, the protein environment induces proton abstraction from 3HF leading to formation of anionic species in the ground state. Fairly high values of anisotropy are observed in the presence of BSA for the tautomer (r = 0.25) as well as anion (r = 0.35) species of 3HF, implying that both the species are located in motion-restricted environments of BSA molecules. Analysis of relevant spectroscopic data leads to the conclusions that two binding sites are involved in BSA-3HF interaction, and the interaction is slightly positively cooperative in nature with a similar binding constant of 1.1 - 1.3 x 10(5) M(-1) for both these sites. Proteins 2001;43:75-81.
Guharay
J
J
Biophysics Division, Saha Institute of Nuclear Physics, 37, Belgachia Road, Calcutta 700 037, India.
Sengupta
B
B
Sengupta
P K
PK
D016880:Anisotropy; D001665:Binding Sites; D004735:Energy Transfer; D005419:Flavonoids / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000032:analysis*; D008962:Models, Theoretical; D011522:Protons; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D013050:Spectrometry, Fluorescence
D005419:Flavonoids; D005456:Fluorescent Dyes; D011522:Protons; D012710:Serum Albumin, Bovine; C041477:3-hydroxyflavone