Bovine serum albumin (BSA)-gold (Au) complexes have been studied in great detail due to their unique red fluorescent (λem = 640 nm) properties, since being initially described by Xie et al. (J. Am. Chem. Soc. 2009, 131, 888-889) as a Au25 nanocluster. We report new findings on these BSA-Au compounds, which were further reducible after synthesis, indicating that these compounds are BSA-cationic Au complexes. We also examined the correlation between pH-induced conformations of BSA and the effect on the resulting red fluorescence to elucidate possible cationic binding sites. The red fluorescence of the BSA-Au complex was associated with only one of five pH-induced isoforms of BSA, the aged (pH > 10) isoform, while the other four BSA conformations (pH < 10) did not result in any red fluorescence. The minimum number of cationic gold per BSA required to yield red fluorescence was less than seven, and internal energy transfer mechanisms could be the reason for the emergence of red fluorescence, based on three-dimensional excitation-emission map measurements. Using various synthesis protocols we determined multiple specific Au binding sites to cysteine disulfide bonds and the N-terminus of the BSA protein. With these findings we present an interpretation of the BSA-Au complex, alternative to the single-site nucleating Au25 nanocluster model.
