For proteins that denature irreversibly, the denaturation is typically triggered by a partial unfolding, followed by a permanent change (e.g., aggregation). The regions that initiate the partial unfolding are named "weak spots". In this work, a molecular dynamics (MD) simulation and data analysis protocol is developed to identify the weak spots of Trichoderma reesei Cel7B, an important endoglucanase in cellulose hydrolysis, through assigning the local melting temperature (Tmp) to individual residue pairs. To test the predicted weak spots, a total of eight disulfide bonds were designed in these regions and all enhanced the enzyme thermostability. The increased stability, quantified by triangle T-50 (which is the T-50 difference between the mutant and the wild type enzyme), is negatively correlated with the MD-predicted T-mp, demonstrating the effectiveness of the protocol and highlighting the importance of the weak spots. Strengthening interactions in these regions proves to be a useful strategy in improving the thermostability of Tr. Cel7B.