Improving Trichoderma reesei Cel7B Thermostability by Targeting the Weak Spots

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• 作者：Shujun Zhang ; Yefei Wang ; Xiangfei Song ; Jingbo Hong ; Yu Zhang ; Lishan Yao
• 刊名：Journal of Chemical Information and Modeling
• 出版年：2014
• 出版时间：October 27, 2014
• 年：2014
• 卷：54
• 期：10
• 页码：2826-2833
• 全文大小：391K
• ISSN：1549-960X

文摘

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 鈥渨eak spots鈥? 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 (T_mp) 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 螖T₅₀ (which is the T₅₀ 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.