Trade-off between Processivity and Hydrolytic Velocity of Cellobiohydrolases at the Surface of Crystalline Cellulose
Trade-off between Processivity and Hydrolytic Velocity of Cellobiohydrolases at the Surface of Crystalline Cellulose
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DOI：
10.1021/ja4119994
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发表时间：
2014-03-26
影响因子：
15
通讯作者：
Samejima, Masahiro
中科院分区：
文献类型：
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作者：
Nakamura, Akihiko;Watanabe, Hiroki;Samejima, Masahiro
由于反应位点的多样性和观察反应的困难，在界面处的多相催化的分析是困难的。纤维素酶对纤维素的酶促水解是固/液界面处的典型非均相反应，并且在聚合物底物上的此类反应的关键参数是持续合成能力，即，在不使酶从底物上脱离的情况下可以发生的催化循环次数。在这项研究中，我们评估了三个密切相关的糖苷水解酶家族7纤维二糖水解酶的丝状真菌在分子水平上的反应，通过高速原子力显微镜，以探讨纤维二糖水解酶的结构与功能的关系，对结晶纤维素。我们发现酶分子在表面上的高运动速度与酶与底物的高解离速率常数相关，这意味着酶与底物之间的弱相互作用。此外，较高的持续合成能力值与覆盖亚位点裂缝的更多环区域相关，这可能意味着较高的结合亲和力。覆盖子位点的环区域导致更强的相互作用，这降低了速度，但增加了持续合成能力。这些结果表明，在加工性纤维素酶之间存在着加工能力和水解速度之间的权衡。
Analysis of heterogeneous catalysis at an interface is difficult because of the variety of reaction sites and the difficulty of observing the reaction. Enzymatic hydrolysis of cellulose by cellulases is a typical heterogeneous reaction at a solid/liquid interface, and a key parameter of such reactions on polymeric substrates is the processivity, i.e., the number of catalytic cycles that can occur without detachment of the enzyme from the substrate. In this study, we evaluated the reactions of three closely related glycoside hydrolase family 7 cellobiohydrolases from filamentous fungi at the molecular level by means of high-speed atomic force microscopy to investigate the structure-function relationship of the cellobiohydrolases on crystalline cellulose. We found that high moving velocity of enzyme molecules on the surface is associated with a high dissociation rate constant from the substrate, which means weak interaction between enzyme and substrate. Moreover, higher values of processivity were associated with more loop regions covering the subsite cleft, which may imply higher binding affinity. Loop regions covering the subsites result in stronger interaction, which decreases the velocity but increases the processivity. These results indicate that there is a trade-off between processivity and hydrolytic velocity among processive cellulases.