Direct Observation of Intermediate State(s) in the Mechanistic Investigation of Domain Specific Protein-Surfactant Interaction

Introduction: Interaction of surfactants with proteins can decipher important information regarding the stability and behavior of proteins. For multi-domain proteins, these interactions vary domain wise and these details are crucial in understanding the contribution of different domains of the protein in its overall activity. Objective: The objective of the present work is to study the interaction of surfactants with domain III of Human Serum Albumin (HSA) and to compare the same with the global interaction. Methods: Interaction of the anionic Sodium Dodecyl Sulphate (SDS) and the Cationic Cetyltrimethylammonium Bromide (CTAB) surfactants with domain III of Human Serum Albumin (HSA) has been studied using 8-Anilino-1-Naphthalene-Sulphonate (ANS) as a fluorescent marker. Circular Dichroism (CD) spectroscopy has been used to study the protein-surfactant interaction for the overall protein. Results: SDS is found to interact sequentially with domain III of HSA having two detectable intermediate states in the binding process. In case of CTAB, we have observed only one intermediate state for its interaction with domain III. Although Quantum yield measurement can reflect the presence of such intermediate state, the overall conformational change of the HSA on addition of surfactants, studied by Circular Dichroism (CD) spectroscopy, and the ANS-Trp distance measurement by FRET could not resolve the presence of such intermediate states. The esterase activity of HSA in presence of different amount of surfactants is also in accordance with our above observation. Conclusion: The interaction of both the surfactants with HSA is found to be sequential in nature. The most important conclusion revealed from our study is that the nature of protein-surfactant interaction is not same throughout the entire protein. Our study reveals that different parts of the multi-domain HSA have different affinity to the surfactant molecules