Naked-Eye Detection of Reversible Protein Folding and Unfolding in Aqueous Solution

• Published in: Journal of chemical education Vol. 94; no. 3; pp. 350 - 355
• Main Authors: Carlson, Tess M, Lam, Kevin W, Lam, Carol W, He, Jimmy Z, Maynard, James H, Cavagnero, Silvia
• Format: Journal Article
• Language: English
• Published: Easton American Chemical Society and Division of Chemical Education, Inc 14-03-2017; American Chemical Society
• Subjects: Aqueous solutions; Biochemistry; Denaturation; Fluorescence; Folding; Learning; Organic chemistry; Protein folding; Proteins; Science education; Serum albumin; First-Year Undergraduate/General; Demonstrations; Upper-Division Undergraduate; Misconceptions/Discrepant Events; Fluorescence Spectroscopy; Proteins/Peptides
• ISSN: 0021-9584; 1938-1328
• DOI: 10.1021/acs.jchemed.6b00507

Students learning general chemistry and biochemistry often have difficulties understanding that proteins routinely exist in both folded and unfolded states, and that protein unfolding is not equivalent to irreversible denaturation or aggregation. To overcome the above misconceptions, we developed a novel demonstration that provides a simple and visually engaging illustration of the reversible interconversion between folded and unfolded protein states. Bovine serum albumin (BSA), a commercially available and inexpensive protein, binds to 8-anilino-1-naphthalenesulfonate (ANS) when folded in solution, and the system fluoresces brightly. Conversely, unfolded BSA (in the presence of ANS) and ANS alone do not fluoresce. Variations in pH and temperature, and addition of destabilizing and stabilizing cosolutes, lead to clear and visually appealing changes in fluorescence, illustrating BSA folding and unfolding. Overall, this novel lecture demonstration aims at overcoming common conceptual challenges, and it demystifies the complex nature of the protein folding/unfolding process by providing a concrete eye-catching example of a generally invisible phenomenon. In addition, this demonstration provides clear evidence for the dramatic effect of pH, temperature, and stabilizing/destabilizing cosolutes on protein conformation.