Crowding-Induced Quenching of Intrinsic Tryptophans of Serum Albumins: A Residue-Level Investigation of Different Conformations

Macromolecular crowding has been known to influence the global conformational landscape of proteins in various ways. However, only a few studies have focused their attention on the local perturbations that can occur in the presence of crowding agents. Here we have used the sensitivity of the tryptophan (Trp) fluorescence of two homologous serum albumins (BSA and HSA) to its (Trp) surroundings to monitor the local changes in the immediate proximity of the intrinsic fluorophore. Using the tool of fluorescence quenching we have shown that the commonly used synthetic macromolecular crowders (e.g., Dextran 6, Dextran 40, Ficoll 70, and PEG 8000) can bring about dramatic conformational modulations in the two proteins. Moreover, the nature of perturbation was observed to be largely dependent on the specific crowding agent used, with Dextran 6 showing the maximum effect while PEG 8000 showed the least. Additionally the extent of local structure modulation was found to be the largest either in the native state of the proteins or under near-native conditions, signifying the important role that the surrounding amino acids play in determining the fluorescence of the Trp residues. Also, surprisingly, although BSA and HSA show ∼76% sequence homology and have almost identical structural disposition in the native state, their individual responses to the crowder-induced perturbation were found to be quite different.