The location of binding of the dyes in human serum albumin influence on dye photochemistry

E. ALARCÓN; A. M. EDWARDS; A. ASPÉE; F.E. MORÁN VIEYRA; C.D. BORSARELLI; E. A. LISSI; DANILO GÓNZALEZ-NILO; HORACIO POBLETE

Salta

Encuentro; 3rd LATIN AMERICAN PROTEIN SOCIETY MEETING; 2010

SAB-CeBEM-LAPSM

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 modeling. 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 behavior. 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 behavior is different to that observed in the external solvent or when it is generated by bound MB.