Circular dichroism spectroscopic detection of ligand binding induced subdomain IB specific structural adjustment of human serum albumin

This work demonstrates for the first time that binding of various compounds within subdomain IB of human serum albumin (HSA) provokes characteristic changes in the near-UV circular dichroism (CD) spectrum of the protein. It can be inferred from the spectroscopic features of difference ellipticity signals and from CD displacement experiments that tyrosine residues located in subdomain IB are the source of the observed spectral alterations. It is proposed that inclusion of some ligand molecules (bile acids, dehydroepiandrosterone sulfate, steroidal terpenes, fatty acids, ibuprofen, and gemfibrozil) into the pocket of subdomain IB disrupts the Tyr138?Tyr161 interhelical π?π stacking interaction, which is reflected in the CD spectrum. This phenomenon can be utilized for the CD detection of subdomain IB specific binding of endo- as well as exogenous agents and to study the drug binding associated local conformational adaptation of the HSA molecule

Bovine Serum Albumin Nanoparticles with Fluorogenic Near-IR-Emitting Squaraine Dyes

Two squaraine (SQ) dyes, N-propanesulfonate-benzothiazolium squaraine (SQ-1) and N-propanesulfonate-benzoindolium squaraine (SQ-2), were synthesized with sulfonate groups to increase water solubility. Both dyes are almost nonfluorescent in aqueous solution with fluorescent quantum yields of 0.03, but exhibited fluorescence enhancement after noncovalently binding with bovine serum albumin (BSA). Upon addition of BSA, the fluorescence intensity increased by ca. a factor of 10, along with a 10-fold extension in the fluorescence lifetime. SQ-1 and SQ-2 interacted with BSA efficiently and appeared to show a preference for binding at site II, which involves combinational effects of electrostatic and hydrophobic interactions. The fluorogenic squaraine dyes were then used to label BSA, forming BSA-based nanoparticles (NPs) through noncovalent binding. The resulting BSA-SQ NPs exhibited enhanced near-IR fluorescence and reduced aggregation of the squaraine moiety. The BSA-SQ NPs were used for cell incubation and bioimaging studies. Confocal fluorescent images were obtained for HCT 116 cells incubated with the BSA-SQ NPs and LysoSensor Green, demonstrating the utility of the NP probes for intracellular imaging. This strategy ovecomes the generally low fluorescence emission of SQ dyes in water and aggregation-reduced fluorescence, providing a versatile strategy for sensing and imaging in biological environments