Poster Presentation 2014 International Biophysics Congress

Thermodynamics of SDS-induced unfolding of helical membrane proteins (#445)

Alvaro A. Recoulat 1 , F Luis Gonzalez-Flecha 1
  1. Department of Biological Chemistry, Facultad de Farmacia y Bioquimica, Universidad de Buenos Aires - CONICET, Buenos Aires, Argentina

Understanding protein folding has been one of the great challenges in biochemistry and molecular biophysics. Although membrane proteins constitute about a third of the proteins encoded in known genomes, folding and stability studies on these proteins have been impaired mainly due to experimental limitations. With only few exceptions1 helical membrane proteins are resistant to chemical denaturation by urea or guanidine hydrochloride. However, ionic detergents, such as SDS, have been efficient denaturant for some of these. The amphipathic nature of detergents combine the possibility of disrupting tertiary contacts and solubilizing the secondary structure elements buried in biological membranes. As chaotropes, SDS binds to both native and denatured proteins, and unfolding is driven by the higher affinity for the denatured state. However, the binding mechanism is different. The driving force contains important favorable entropic contributions, making the denaturing effect evident at millimolar concentrations. Besides, hydrophobic clusters of SDS molecules are able to induce helical structure in amphipathic non-structured chains. Thus, the final denatured state has high secondary structure content. Thermodynamic characterization of membrane protein stability requires experimental conditions where unfolding can be reversibly achieved2. When this is possible, equilibrium thermodynamics provides the framework for determining the Gibbs free-energy change for transfer the membrane protein from an ideal one molar solution of the pure folded protein in water to an ideal one molar aqueous solution of the pure “unfolded” protein, which is the accepted form to quantify the structural stability of a protein. Given the differences between the unfolded states induced by urea (or guanidine hydrochloride) and that formed when SDS is used as denaturant, comparison of the unfolding thermodynamic parameters cannot be directly performed although both quantities are extrapolated to zero denaturant concentration. 

With Grants from UBACyT (20020100100048) and ANPCyT (PICT2010-01741)

  1. Roman EA. Argüello JM, González Flecha FL. Reversible Unfolding of a Thermophilic Membrane Protein in Phospholipid/Detergent Mixed Micelles. J Mol Biol 2010, 397, 550-559
  2. Roman EA, González Flecha FL. Kinetics and Thermodynamics of Membrane Protein Folding. Biomolecules 2014, 4, 354-373.