Colloidal protein stability and surface coverage by trehalose: bridging molecular dynamics and experimental work

The aim of this study was to assess to what extent a simulated trehalose monolayer can predict the minimum molar trehalose to protein ratios required to preserve the colloidal stability of lyophilized lactate dehydrogenase, catalase, and β-galactosidase. We employed an iterative molecular dynamics (MD) approach to quantify the number of trehalose molecules required to saturate the protein's accessible surface area. These in-silico values were compared to experimentally determined colloidal stability thresholds (tetramer recovery) in lyophilized formulations, defined as the minimum molar ratio required to maintain hydrodynamic volume, as assessed by size exclusion chromatography following storage at 30°C and 43% relative humidity for a week. A quantitative, protein-specific correlation was observed between the simulated surface coverage and the experimental colloidal stability thresholds. This result shows that MD simulations can serve as a powerful pre-formulation tool for predicting minimum excipient requirements and provides a foundation for a more rational approach to protein stabilization.