Processive Degradation of Crystalline Cellulose by a Multimodular Endoglucanase via a Wirewalking Mode
Processive Degradation of Crystalline Cellulose by a Multimodular Endoglucanase via a Wirewalking Mode
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多模块内切葡聚糖酶通过走丝模式持续降解结晶纤维素
DOI：
10.1021/acs.biomac.8b00340
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发表时间：
2018
影响因子：
6.2
通讯作者：
Wang LS
中科院分区：
文献类型：
--
作者：
Zhang Kun-Di;Wang Ye-Fei;Tan Fang-Cheng;Yao Li-Shan;Li Fu-Li;Li Wen;Wang Lu-Shan;Zheng Yan-Lin;Bayer Edward A.;Ma Xiao-Qing;Li FL;Wang LS
纤维素酶对结晶纤维素的过程性水解是木质纤维素降解的关键步骤。经典的里氏木霉外切葡聚糖酶TrCel 7A具有封闭的活性位点隧道，通过用纤维素链穿过隧道来开始每次进行性运行。环区是隧道构象所必需的，导致真菌外切葡聚糖酶的弱热稳定性。然而，来自嗜热细菌纤维梭菌的内切葡聚糖酶CcCel 9A包含具有开放裂缝的糖苷水解酶（GH）家族9模块和五个碳水化合物结合模块（CBM），并且水解结晶纤维素原。CcCel 9A和其他类似的GH 9酶如何结合到结晶纤维素的光滑表面以实现持续合成能力仍然是未知的。我们的研究结果表明，C-末端的CBM 3b和三个CBMX 2增强生产吸附纤维素，而CBM 3c相邻的GH 9是紧密结合到11个葡萄糖基单位，从而延长催化裂缝17个亚位点，这有利于decomponization通过形成一个supermodular的结合表面。在开放裂缝中，底物结合亚位点和葡萄糖环之间的强相互作用力使氢键断裂和提取单个纤维素链成为可能。此外，子站点-4能够将链拉到它喜欢的位置。纤维四糖作为初始产物从开放裂缝释放以实现高持续合成能力，其通过内切葡聚糖酶的催化裂缝进一步水解为纤维三糖、纤维二糖和葡萄糖。在此基础上，提出了内切葡聚糖酶连续降解结晶纤维素的走丝模式，为工业纤维素酶的合理设计提供了思路。
Processive hydrolysis of crystalline cellulose by cellulases is a critical step for lignocellulose deconstruction. The classicTrichoderma reeseiexoglucanaseTrCel7A, which has a closed active-site tunnel, starts each processive run by threading the tunnel with a cellulose chain. Loop regions are necessary for tunnel conformation, resulting in weak thermostability of fungal exoglucanases. However, endoglucanaseCcCel9A, from the thermophilic bacteriumClostridium cellulosi, comprises a glycoside hydrolase (GH) family 9 module with an open cleft and five carbohydrate-binding modules (CBMs) and hydrolyzes crystalline cellulose processively. HowCcCel9A and other similar GH9 enzymes bind to the smooth surface of crystalline cellulose to achieve processivity is still unknown. Our results demonstrate that the C-terminal CBM3b and three CBMX2s enhance productive adsorption to cellulose, while the CBM3c adjacent to the GH9 is tightly bound to 11 glucosyl units, thereby extending the catalytic cleft to 17 subsites, which facilitates decrystallization by forming a supramodular binding surface. In the open cleft, the strong interaction forces between substrate-binding subsites and glucosyl rings enable cleavage of the hydrogen bonds and extraction of a single cellulose chain. In addition, subsite −4 is capable of drawing the chain to its favored location. Cellotetraose is released from the open cleft as the initial product to achieve high processivity, which is further hydrolyzed to cellotriose, cellobiose and glucose by the catalytic cleft of the endoglucanase. On this basis, we propose a wirewalking mode for processive degradation of crystalline cellulose by an endoglucanase, which provides insights for rational design of industrial cellulases.