Xia_2026_Biochim.Biophys.Acta.Proteins.Proteom_1875_141186

The use of enzymes in moist-solid reaction mixtures with gentle mechanical mixing (mechanoenzymology) is an emerging technology of high promise for the development of more sustainable processes. For example, this strategy enables the depolymerization of highly crystalline polyethylene terephthalate (PET) without the need for a dedicated thermal amorphization pretreatment or pH control, while also generating a clean product and minimal wastewater. Although these low water conditions have been reported to enhance the activity of several enzymes, the approach remains underexplored and poorly understood. Herein, the hydrolysis of PET catalyzed by the commercial cutinase Novozym 51,032 (abbreviated HiC) is used as a model system to study enzyme behavior in moist solids. Since fractal kinetics can describe reactions in heterogeneous systems, we report here the temperature-dependent behavior of fractal kinetics for this reaction, as well as the first estimation of activation energy and thermodynamic parameters for an enzymatic reaction in such low-water systems. Overall, this work not only provides more mechanistic insights into this reaction but also serves as a template for estimating energy contributions of other reactions in moist-solid mixtures.