Practice question

Accurate molecular weight estimation by SDS-PAGE requires both anionic detergent and reducing thiol for complete linearization. Sodium dodecyl sulfate disrupts non-covalent interactions and imparts uniform negative charge but cannot cleave covalent disulfide linkages between cysteine residues, which constrain polypeptide loops and maintain compact domains. Addition of beta-mercaptoethanol or dithiothreitol reduces disulfides to free sulfhydryls, fully extending chain. Without reducing agent, proteins remain denatured yet partially constrained with intact disulfides, resulting in anomalously fast or slow migration, incomplete unfolding and inaccurate mass estimation and possible multimer persistence.