Abstract
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This study was performed to check the effect of temperature on the refolding of acid-denatured horse heart cytochrome c (h-cyt c) at pH 2.0 under the influence of anionic surface active ionic liquid (SAIL) 1-butyl-3-methylimidazolium octyl sulfate ([C4mim][C8OSO3]), various techniques were performed viz. UV–vis, steady-state fluorescence, time-resolved fluorescence, circular dichroism (CD), FT-IR and dynamic light scattering. We found SAIL, [C4mim][C8OSO3] upon addition to the h-cyt c in unfolded state at pH 2.0, stabilized the molten globule (MG), which acts as an intermediate during the protein folding pathway. The investigation of the refolding of h-cyt cin an unfolded state with [C4mim][C8OSO3] shows a surfactant like behavior and stabilizes the MG-like state with the help of electrostatic and hydrophobic interactions. The creation of an MG-like state reveals the presence of a regular structure formed in the presence of SAIL [C4mim][C8OSO3]. The thermodynamic parameters of the refolding of h-cyt c by [C4mim][C8OSO3] were also quantified at various temperatures (25 °C–40 °C). The m value calculated for refolded states of h-cyt c by [C4mim][C8OSO3] shows a noticeable difference. The lower values of m (at higher temperatures) suggest less efficient refolding. This shows the refolding ability of the [C4mim][C8OSO3] decreases with increasing the temperature. Moreover, the anionic SAILs were efficiently utilized in the protein renaturation studies and could be effectively used to solve problems of misfolding and aggregation of proteins.