Kinetics of Fluorophore Formation in Bovine Serum Albumin–Gold Complexes

We revisit the prevailing hypothesis that the red fluorophore (λ_em = 640 nm) in the bovine serum albumin (BSA)–gold (Au) compound is a Au₂₅ nanocluster. To examine the hypothesis, we investigated the kinetics of Au binding in this compound. In addition to the specific Au­(III) binding sites in BSA, we found a significant degree of nonspecific Au­(III) binding on the BSA surface. Time-course of the emergence of the red fluorescence was measured in detail for a range of pH, temperature, and concentration of Au­(III) with respect to BSA. The red fluorophore formation was a slow yet dynamic process, which was consistent with the pH-induced equilibrium transition in the conformation of BSA. Notably, the kinetic rate of the fluorophore formation was not strongly dependent on the concentration of Au­(III). Incorporating the existence of multiple specific and nonspecific binding sites, we propose a new model of the red fluorophore formation mechanism based on Langmuir-type adsorption of Au to BSA, as an alternative to the single-site nucleation model of Au₂₅ nanoclusters.