Photophysics and photochemistry of dyes bound to human serum albumin are determined by the dye localization

Abstract

The photophysics and photochemistry of rose bengal (RB) and methylene blue (MB) bound to human serum albumin (HSA) have been investigated under a variety of experimental conditions. Distribution of the dyes between the external solvent and the protein has been estimated by physical separation and fluorescence measurements. The main localization of protein-bound dye molecules was estimated by the intrinsic fluorescence quenching, displacement of fluorescent probes bound to specific protein sites, and by docking modelling. All the data indicate that, at low occupation numbers, RB binds strongly to the HSA site I, while MB localizes predominantly in the protein binding site II. This different localization explains the observed differences in the dyes' photochemical behaviour. In particular, the environment provided by site I is less polar and considerably less accessible to oxygen. The localization of RB in site I also leads to an efficient quenching of the intrinsic protein fluorescence (ascribed to the nearby Trp residue) and the generation of intra-protein singlet oxygen, whose behaviour is different to that observed in the external solvent or when it is generated by bound MB.

Effect of dye localization and self-interactions on the photosensitized generation of singlet oxygen by rose bengal bound to bovine serum albumin

“…These results suggest that in both albumins the dye bounds into a hydrophobic protein pocket with similar nanoenvironmental properties, but with lower affinity for BSA. In the case of HSA, dansyl-probes displacement by RB experiments demonstrated that the dye preferably binds to the hydrophobic subdomain IIA (or Sudlow's site I) of the protein [12,13], as it is generally accepted for bulky heterocyclic anions [14]. Considering that BSA shares %76% of the amino acidic sequence homology with HSA [14], it can be also expected the binding of RB into site I of BSA.…”

“…Triplet (U T ) and bleaching (U b ) quantum yields of RB in BSA solutions were calculated by comparison of the respective transient absorption changes at t = 0 (DA 0 ) at each maximum divided by the ground state absorption of the solution at the laser excitation wavelength with that observed for the dye alone in buffer solution, assuming the same extinction coefficient of the transient species without and with protein [12,13].…”

“…On the other hand, as a different strategy to obtain efficient biocompatible 1 O 2 -photosensitizer systems, we have characterized the photophysical and photochemical properties of well-known Type II artificial sensitizers such as zinc phthalocyanine (ZnPc) [6], rose bengal (RB) [12], and methylene blue (MB) [13], non-covalently bound to both bovine and human serum albumins (BSA and HSA), considering the large concentration of these proteins in blood and their function in the transport and delivery of small drugs and bioactive molecules [14]. However, despite of the moderate efficiency of generation of 1 O 2 of these supramolecular photosensitizer assemblies, the photophysical and photosensitizing properties of the dye-albumin adducts were mainly dependent on the type of binding-site in the albumin, molecular oxygen accessibility to the dye binding location, and also of the average number of sensitizer molecules per protein (n), being each adduct system practically a particular case of study.…”

Interaction of 1-pyrene sulfonic acid sodium salt with human serum albumin

“…This structure provides two principal binding sites for aromatic and heterocyclic molecules, known as Sudlow's sites 1 and 2 [10][11][12] which are located in subdomains IIA and IIIA, respectively; although some other low-affinity sites also exist [13][14][15].…”

Synthesis and Physicochemical Characterization of Biodegradable Star-Shaped Poly Lactide-Co-Glycolide-<i>β</i>-Cyclodextrin Copolymer Nanoparticles Containing Albumin

“…So serum albumin is in use as a model for studying drug-protein interaction in vitro. Many drugs and other bioactive small molecules bind reversibly to albumin and other serum components then utility as carriers [26].…”