Molecular Basis for the Solvation and Reconstructive Denaturation of Proteins by Detergents

Abstract

Detergents are widely used for the biochemical and structural study of proteins. Non-ionic and zwitterionic detergents are used as membrane mimetics, where they solvate the hydrophobic regions of integral membrane proteins. In contrast, ionic detergents such as anionic sodium dodecyl sulfate (SDS) and cationic lauryl trimethylammonium chloride (LTAC) are strong protein denaturants that unfold both soluble and membrane proteins. Not only does the SDS-unfolded state have high α-helix content, but SDS also induces α-helix formation irrespective of the intrinsic native secondary structure, a process known as “reconstructive denaturation.” Although this latter phenomenon underpins the ubiquitous technique SDS-PAGE, the mechanism of SDS denaturation and the molecular nature of the SDS denatured state are not known. We use a combined biophysical and computational approach to elucidate the molecular basis of protein denaturation by ionic detergents, with a special focus on the mechanism of reconstructive denaturation by SDS. Specifically, biophysical techniques, including CD and ITC, are used to study the interaction of a set of detergents with model systems that build in complexity from model peptides to full proteins. In the first case, peptide sequence design is used to explore the specific features determining interactions with ionic detergents, while in the latter, chemical modification of residue side chains is used to explore the determinants of detergent interactions in the complex background of a biological protein sequence. In parallel with the biophysical studies, molecular dynamics simulations of the same systems are used to provide atomic resolution detail of the results from the biophysical experiments, and to determine the modes of detergent micelle-protein interaction. Together, these results suggest a consistent mechanism for the reconstructive denaturation phenomenon, for sequence based specificity of ionic detergent interactions, and ultimately, for SDS's universal protein denaturing action.