Secondary structural changes in guanidinium hydrochloride denatured mammalian serum albumins and protective effect of small amounts of cationic gemini surfactant pentanediyl-a,x- bis(cetyldimethylammonium bromide) and methyl-b-cyclodextrin: A spectroscopic study Nuzhat Gull a,⇑ , Javed Masood Khan b , Mohd Ishtikhar b , Atiyatul Qadeer b , Rehan Ajmal Khan b , Mudasir Gul c , Rizwan Hasan Khan b,⇑ a Department of Chemistry, Govt. Degree College for Women, M. A. Road, Srinagar, India b Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh 202 002, India c Jawahar Lal Nehru Medical College and Hospital, Aligarh Muslim University, Aligarh 202 002, India article info Article history: Received 3 September 2014 Accepted 10 October 2014 Available online 20 October 2014 Keywords: Sheep serum albumin Rabbit serum albumin Porcine serum albumin Pentanediyl-a,x- bis(cetyldimethylammonium bromide) Circular dichroism and dynamic light scattering abstract In the present study the cationic gemini surfactant assisted refolding of guanidinium hydrochloride (GdCl) denatured mammalian serum albumins viz. sheep serum albumin (SSA), rat serum albumin (RSA) and porcine serum albumin (PSA) using a combination of cationic gemini surfactants, penta- nediyl-a,x-bis(cetyldimethylammonium bromide) (C 16 H 33 (CH 3 ) 2 N + –(CH 2 ) 5 –N + (CH 3 ) 2 C 16 H 33 )2Br  desig- nated as G5 and methyl-b-cyclodextrin in the artificial chaperone assisted two step method, is attempted. The studies were carried out in an aqueous medium (pH 7.4) using dynamic light scattering (DLS), circu- lar dichroism (CD), and fluorescence spectroscopy. A perusal of DLS data indicates that against the native hydrodynamic radius (R h ) of 4.3 nm in SSA, 3.9 nm in PSA and 3.5 nm in RSA, the R h of the said proteins, when refolding is attempted by simple dilution, increases to 21.7 nm, 36.6 nm and 37.2 nm, respectively. Hydrodynamic radii very near to the native protein, i.e., 4.0 nm, 4.1 nm and 4.4 nm for RSA, PSA and SSA respectively, is obtained on the sequential addition of G5 and methyl-b-cyclodextrin to the denatured protein. Circular dichroism studies corroborate with the DLS data. The results obtained from the multi- technique approach are ascribed to the presence of two charged head-groups and two hydrocarbon tails in the gemini surfactants resulting in a very strong electrostatic and hydrophobic interactions. Based on the present study it is suggested that the gemini surfactants may be utilized in the protein refolding stud- ies and thus may address one of the most pressing demand of biotechnology industry for the develop- ment of efficient and inexpensive folding aides. Ó 2014 Elsevier Inc. All rights reserved. 1. Introduction Recombinant DNA technology has made available several sim- ple techniques that could have made accessible unlimited and inexpensive sources of otherwise rare proteins. However, such pro- teins are often aggregated in the form of inactive inclusion bodies [1,2]. The inclusion bodies can be solubilized in concentrated chaotropes but it results in a protein devoid of its native conforma- tion. As the proteins must fold into its characteristic and functional three dimensional structures in order to function [3,4], the dena- turant concentration has to be lowered to allow the formation of the native protein conformation. When proper conditions for refolding are identified the correct refolding process competes, often in disadvantage, with misfolding and aggregation [5–7]. Pro- tein misfolding and aggregation pose a serious problem in the industrial process of producing the recombinant proteins [8,9]. In order to develop efficient folding processes the competition between folding and aggregation in ‘in vitro’ proteins has to be understood. Aggregation may be due to the association of hydro- phobic surfaces that are exposed during the refolding process http://dx.doi.org/10.1016/j.jcis.2014.10.012 0021-9797/Ó 2014 Elsevier Inc. All rights reserved. Abbreviations: SSA, sheep serum albumin; RSA, rabbit serum albumin; PSA, porcine serum albumin; G5, pentanediyl-a,x-bis(cetyldimethylammonium bromide); CD, circular dichroism; DLS, dynamic light scattering. ⇑ Corresponding authors. E-mail addresses: nuzhatwahid@gmail.com (N. Gull), rizwanhkhan@hotmail. com (R.H. Khan). Journal of Colloid and Interface Science 439 (2015) 170–176 Contents lists available at ScienceDirect Journal of Colloid and Interface Science www.elsevier.com/locate/jcis