A complete picture of protein unfolding and refolding in surfactants

CIC nanoGUNE Seminars

Daniel Otzen, Aarhus University
nanoGUNE seminar room, Tolosa Hiribidea 76, Donostia - San Sebastian
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A complete picture of protein unfolding and refolding in surfactants Systems containing proteins and surfactants are of broad interest, scientifically as well as technologically, owing to their widespread application in detergency and various biotechnologies. It is by now well- established (by our group and others) that the anionic surfactant SDS denatures most globular proteins; furthermore, the end-point of the unfolding reaction is a complex consisting of nearly intact SDS micelles with the unfolded proteins wrapped around the micelles. The proteins' alpha-helical secondary structure is either preserved from the native state or (more often) induced by the interaction with the micelle. Recently, we have also shown that addition of non-ionic surfactant to solutions with SDS-protein complexes leads to refolding of the proteins and to regain of secondary structure also for beta-sheet proteins. While numerous studies, several of which from our own groups, have investigated the unfolding and refolding processes at the level of the protein, there is a major gap in our understanding: we simply do not know how the surfactants and proteins cooperate or "dance together" to facilitate these processes. The key question is as follows: What are the steps by which the protein encounters the surfactant micelle (and vice versa) and how do they both rearrange in response to the other component in order to lead to a new state, either an unfolded protein-surfactant complex or - in the case of refolding - a protein liberated from the "shackles" of the original denaturing anionic micelle? We present the first work which studies the kinetics of unfolding and refolding of a protein as it interacts with surfactants, both from the perspective of the protein and the surfactant. The key to this new understanding is to exploit synchrotron radiation sources to record stopped- flow Small Angle X-ray Scattering spectra at low-ms resolution of the evolving protein-surfactant complexes. By combining this with stopped-flow circular dichroism and tryptophan fluorescence we have for the first time obtained a detailed picture of the protein and surfactant structures involved in each step of the unfolding and refolding process. The combination of the techniques has given unique information on the various steps, in the time scales and on the involved intermediate structure. Our work reveals an intriguing sequence of events: unfolding involves redistribution of the protein around the micelle while refolding involves parallel detachment and refolding steps. **Host:** A. Bittner