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A complete picture of protein unfolding and refolding in surfactants.

ABSTRACT: Interactions between proteins and surfactants are of relevance in many applications including food, washing powder formulations, and drug formulation. The anionic surfactant sodium dodecyl sulfate (SDS) is known to unfold globular proteins, while the non-ionic surfactant octaethyleneglycol monododecyl ether (C₁₂E₈) can be used to refold proteins from their SDS-denatured state. While unfolding have been studied in detail at the protein level, a complete picture of the interplay between protein and surfactant in these processes is lacking. This gap in our knowledge is addressed in the current work, using the β-sheet-rich globular protein β-lactoglobulin (bLG). We combined stopped-flow time-resolved SAXS, fluorescence, and circular dichroism, respectively, to provide an unprecedented in-depth picture of the different steps involved in both protein unfolding and refolding in the presence of SDS and C₁₂E₈. During unfolding, core-shell bLG-SDS complexes were formed within ∼10 ms. This involved an initial rapid process where protein and SDS formed aggregates, followed by two slower processes, where the complexes first disaggregated into single protein structures situated asymmetrically on the SDS micelles, followed by isotropic redistribution of the protein. Refolding kinetics (>100 s) were slower than unfolding (<30 s), and involved rearrangements within the mixing deadtime (∼5 ms) and transient accumulation of unfolded monomeric protein, differing in structure from the original bLG-SDS structure. Refolding of bLG involved two steps: extraction of most of the SDS from the complexes followed by protein refolding. These results reveal that surfactant-mediated unfolding and refolding of proteins are complex processes with rearrangements occurring on time scales from sub-milliseconds to minutes.

SUBMITTER: Pedersen JN

PROVIDER: S-EPMC8145811 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The interaction of protein and surfactant yields protein-surfactant complexes which have a wide range of applications in the cosmetics, foods, and pharmaceutical industries among others. Ionic and nonionic surfactants are known to interact differently with the protein. The interplay of electrostatic and hydrophobic interactions governs the resultant structure of protein-surfactant complexes. The present study enlightens the paramount role of the hydrophobic interaction, tuned by the hydrophobic tail length of ionic surfactants, in the unfolding of anionic bovine serum albumin (BSA) protein. The unfolding of BSA in the presence of four different tail-length cationic surfactants, that is, C10TAB, C12TAB, C14TAB, and C16TAB, has been investigated by small-angle neutron scattering and dynamic light scattering. All cationic surfactants unfold the protein at a certain concentration range. The propensity of protein unfolding increases with increasing the hydrophobic tail length. The denatured structure of BSA upon addition of cationic surfactants is characterized by the random flight model representing a beads-on-a-string chain-like complex. The unfolded protein binds the surfactant micelles in the protein-surfactant cluster. The micelles get elongated with the increasing concentration of cationic surfactants, whereas the number of micelles per cluster is decreased. In the final stage, the protein-surfactant cluster merges to one large micelle with unfolded protein wrapping the micelle surface. The pathway of protein unfolding is described in terms of the changes in the micellar size, the number of micelles formed per cluster, the separation between the micelles in the cluster, the aggregation number of micelles, and the number of proteins per cluster. The protein-surfactant interaction is further examined in the presence of a nonionic surfactant, that is, C12E10. The nonionic surfactant significantly suppresses the interaction of BSA protein with ionic surfactants by forming mixed micelles. As a result of the mixed micelles formation by ionic-nonionic surfactants, the ionic surfactant moves out from the unfolded BSA protein, and this enables the protein to refold back to its native structure. The propensity of mixed micelle-driven refolding of proteins is significantly changed with changing the tail length of the ionic surfactant.

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A complete picture of protein unfolding and refolding in surfactants.

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