Probing the binding mode and interactions of proteinase K and glutathione: molecular simulation and experiments

Abstract

Proteinase K, a serine protease from Tritirachium album Limber, is crucial in research due to its potent proteolytic activity, which relies on conformational stability and substrate affinity. Glutathione (GSH), an essential intracellular antioxidant, regulates various physiological processes by interacting with proteins, influencing their stability and function. Despite the importance of both proteinase K and GSH, their potential interaction remains unexplored. Understanding this interaction could uncover new regulatory mechanisms affecting proteinase K, with significant implications for research and therapeutic applications. In this study, we systematically investigated the binding of GSH to proteinase K using a comprehensive approach in which theoretical and experimental methods mutually validate each other. Molecular docking determined the binding mode and the interaction mechanism of proteinase K and GSH. Molecular dynamics (MD) simulations revealed that GSH binding significantly improved the stability of proteinase K, affirming the binding process was spontaneous, with hydrogen bonds and van der Waals forces emerging as the predominant contributors throughout the interaction. At the same time, the fluorescence spectrum and circular dichroism spectrum confirmed the interaction mechanism between GSH and proteinase K, as well as the conformational changes of proteinase K induced by GSH binding. We believe this study could offer valuable insights for future research into the structure and binding dynamics of other protein–ligand complexes under physiological conditions.