Refolding of bovine serum albumin via artificial chaperone protocol using gemini surfactants

Surfactants prevent the irreversible aggregation of partially refolded proteins, and they are also known to assist in protein refolding. A novel approach to protein refolding that utilizes a pair of low molecular weight folding assistants, a detergent and cyclodextrin, was proposed by Rozema and Gellman (D. Rozema, S.H. Gellman, J. Am. Chem. Soc. 117 (1995) 2373). We report the refolding of bovine serum albumin (BSA) assisted by these artificial chaperones, utilizing gemini surfactants for the first time. A combination of cationic gemini surfactants, bis(cetyldimethylammonium)pentane dibromide (C16H33(CH3)2N+–(CH2)5–N+(CH3)2C16H33·2Br− designated as G5 and bis(cetyldimethylammonium)hexane dibromide (C16H33(CH3)2N+–(CH2)6–N+(CH3)2C16H33·2Br− designated as G6 and cyclodextrins, was used to refold guanidinium chloride (GdCl) denatured BSA in the artificial chaperone assisted two step method. The single chain cationic surfactant cetyltrimethylammonium bromide (CTAB) was used for comparative studies. The studies were carried out in an aqueous medium at pH 7.0 using circular dichroism, dynamic light scattering and ANS binding studies. The denatured BSA was found to get refolded by very small concentrations of gemini surfactant at which the single chain counterpart was found to be ineffective. Different from the single chain surfactant, the gemini surfactants exhibit much stronger electrostatic and hydrophobic interactions with the protein and are thus effective at much lower concentrations. Based on the present study it is expected that gemini surfactants may prove useful in the protein refolding operations and may thus be effectively employed to circumvent the problem of misfolding and aggregation.

Graphical abstractThe present finding shows the excellent efficacy of gemini surfactants as capturing agents during artificial chaperone protocol for achieving refolding of BSA.Figure optionsDownload full-size imageDownload high-quality image (121 K)Download as PowerPoint slideHighlights► We report refolding of BSA through artificial chaperone protocol using gemini surfactants. ► The studies were performed using circular dichroism, DLS and fluorescence studies. ► Results show geminis’ to be more useful in the protein refolding than conventional surfactants.