A disulfide redox switch mechanism regulates glycoside hydrolase function

Disulfide bond is a key post-translational modification involved in protein folding, stability, and functional regulation. While the structural role of disulfide bonds has been extensively studied in protein chemistry, their potential involvement in regulating catalytic activity of carbohydrate-active enzymes through redox switch mechanisms remains an untapped realm. In this work, we demonstrate that a glycoside hydrolase from the large family GH2 has its catalytic activity regulated via an intramolecular disulfide bond, adapting dynamically to redox fluctuations in its environment. The enzyme is inactive in its oxidized state, becoming active when reduced through a fully reversible process. Under oxidative conditions, multiple crystallographic structures showed that the disulfide bond formation induces a massive structural disorder in the active site, disrupting the substrate binding site and remarkably the acidic catalytic residues configuration. Conversely, high-resolution cryo-EM structure of the active (reduced) state revealed a notable well-structured active site with catalytic residues properly positioned for a classical Koshland retaining mechanism. This reversible order-disorder mechanism based on disulfide switch to regulate catalytic activity broadens our understanding regarding redox systems involved in carbohydrate breakdown and metabolism, which have implications for biotechnology and microbial biology.