Extracellular enzymes are mainly responsible for depolymerizing soil organic matter (SOM) in terrestrial ecosystems, and soil minerals are known to affect enzyme activity. However, the mechanisms and the effects of mineral-enzyme interactions on enzymatic degradation of organic matter remain poorly understood. In this study, we examined the adsorption of fungal β-glucosidase enzyme on minerals and time-dependent changes of enzymatic reactivity, measured by the degradation of two organic substrates (i.e., cellobiose and indican) under both cold (4 °C) and warm (20 and 30 °C) conditions. Hematite, kaolinite, and montmorillonite were used, to represent three common soil minerals with distinctly different surface charges and characteristics. β-glucosidase was found to sorb more strongly onto hematite and kaolinite than montmorillonite. All three minerals inhibited enzyme degradation of cellobiose and indican, likely due to the inactivation or hindrance of enzyme active sites. The mineral-bound β-glucosidase retained its specificity for organic substrate degradation, and increasing temperature from 4 to 30 °C enhanced the degradation rates by 2–4 fold for indican and 5–9 fold for cellobiose. These results indicate that enzyme adsorption, mineral type, temperature, and organic substrate specificity are important factors influencing enzymatic reactivity and thus have important implications in further understanding and modeling complex enzyme-facilitated SOM transformations in terrestrial ecosystems.