Human serum albumin (HSA), being the most abundant carrier protein in blood and a modern day clinical tool for drug delivery, attracts high attention among biologists. Hence, its unfolding/refolding strategies and exogenous/endogenous ligand binding preference are of immense use in therapeutics and clinical biochemistry. Among its fellow proteins albumin is known to carry almost every small molecule. Thus, it is a potential contender for being a molecular cargo/or nanovehicle for clinical, biophysical and industrial purposes. Nonetheless, its structure and function are largely regulated by various chemical and physical factors to accommodate HSA to its functional purpose. This multifunctional protein also possesses enzymatic properties which may be used to convert prodrugs to active therapeutics. This review aims to highlight current overview on the binding strategies of protein to various ligands that may be expected to lead to significant clinical applications.

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Little work has been done to understand the folding profiles of multi-domain proteins at alkaline conditions. We have found the formation of a molten globule-like state in bovine serum albumin at pH 11.2 with the help of spectroscopic techniques; like far and near ultra-violet circular dichroism, intrinsic and extrinsic fluorescence spectroscopy. Interestingly, this state has features similar to the acid-denatured state of human serum albumin at pH 2.0 reported by Muzammil et al. (Eur J Biochem 266:26-32, 1999). This state has also shown significant increase in 8-anilino-1-naphthalene-sulfonate (ANS) binding in compare to the native state. At pH 13.0, the protein seems to acquire a state very close to 6 M guanidinium hydrochloride (GuHCl) denatured one. But, reversibility study shows it can regain nearly 40% of its native secondary structure. On the contrary, tertiary contacts have disrupted irreversibly. It seems, withdrawal of electrostatic repulsion leave room for local interactions, but disrupted tertiary contacts fail to regain their original states.

The interactions among bovine, rabbit, and porcine serum albumins and single-chain cationic surfactant cetyltrimethylammonium bromide (CTAB) versus its gemini counterpart (designated as G4) have been studied. The studies were carried out in an aqueous medium at pH 7.0 using UV, intrinsic and extrinsic fluorescence spectroscopy, and far-UV circular dichroism techniques. The results indicate that compared to CTAB, G4 interacts strongly with the serum albumins, resulting in a significantly larger unfolding or decrease in alpha-helical content as reflected by the significantly larger decrease in ellipticity in the far-UV range. Unlike CTAB, a remarkable increase in the alpha-helical content of BSA at 625 microM G4 and at 250 microM G4 for RSA and PSA is observed. The appearance of conformational changes and saturation points in the proteins occurs at considerably lower [G4] compared to [CTAB]. The results obtained from the multi-technique approach are ascribed to the stronger forces in G4 owing to the presence of two charged headgroups and two hydrocarbon tails. Keeping the results in view, it is suggested that the gemini surfactants may be effectively used in the renaturation of proteins produced in genetically engineered cells via the artificial chaperone protocol and may also prove useful in drug delivery as solubilizing agents to recover proteins from insoluble inclusion bodies.

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