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首页> 外文期刊>Applied Microbiology >Interplay between Clostridium thermocellum Family 48 and Family 9 Cellulases in Cellulosomal versus Noncellulosomal States
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Interplay between Clostridium thermocellum Family 48 and Family 9 Cellulases in Cellulosomal versus Noncellulosomal States

机译：热纤梭菌家族48和9纤维素酶在纤维素和非纤维素状态下的相互作用。

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The anaerobic, thermophilic cellulolytic bacterium Clostridium thermocellum is known for its elaborate cellulosome complex, but it also produces a separate free cellulase system. Among the free enzymes, the noncellulosomal enzyme Cel9I is a processive endoglucanase whose sequence and architecture are very similar to those of the cellulosomal enzyme Cel9R; likewise, the noncellulosomal exoglucanase Cel48Y is analogous to the principal cellulosomal enzyme Cel48S. In this study we used the designer cellulosome approach to examine the interplay of prominent cellulosomal and noncellulosomal cellulases from C. thermocellum . Toward this end, we converted the cellulosomal enzymes to noncellulosomal chimeras by swapping the dockerin module of the cellulosomal enzymes with a carbohydrate-binding module from the free enzyme analogues and vice versa . This enabled us to study the importance of the targeting effect of the free enzymes due to their carbohydrate-binding module and the proximity effect for cellulases on the designer cellulosome. C. thermocellum is the only cellulosome-producing bacterium known to express two different glycoside hydrolase family 48 enzymes and thus the only bacterial system that can currently be used for such studies. The different activities with crystalline cellulose were examined, and the results demonstrated that the individual chimeric cellulases were essentially equivalent to the corresponding wild-type analogues. The wild-type cellulases displayed a synergism of about 1.5-fold; the cellulosomal pair acted synergistically when they were converted into free enzymes, whereas the free enzymes acted synergistically mainly in the wild-type state. The targeting effect was found to be the major factor responsible for the elevated activity observed for these specific enzyme combinations, whereas the proximity effect appeared to play a negligible role.
机译：厌氧，高温纤维素分解细菌热纤梭菌（Clostridium thermocellum）以其复杂的纤维素体复合物而闻名，但它也产生了一个独立的游离纤维素酶系统。在游离酶中，非纤维素酶Cel9I是一种进行性内切葡聚糖酶，其序列和结构与纤维素酶Cel9R的非常相似。同样地，非纤维素外切葡聚糖酶Cel48Y类似于主要的纤维素酶Cel48S。在这项研究中，我们使用了设计的纤维素体方法研究了C. thermocellum的突出的纤维素和非纤维素纤维素酶的相互作用。为此，我们通过将纤维素酶的dockerin模块与游离酶类似物的碳水化合物结合模块交换，将纤维素酶转化为非纤维素嵌合体。这使我们能够研究由于游离酶的碳水化合物结合模块而产生的靶向作用的重要性，以及纤维素酶对设计纤维素团的邻近效应。热纤梭菌是已知唯一表达两种不同糖苷水解酶家族48酶的产纤维素酶的细菌，因此是目前可用于此类研究的唯一细菌系统。检查了结晶纤维素的不同活性，结果表明各个嵌合纤维素酶基本上等同于相应的野生型类似物。野生型纤维素酶显示出约1.5倍的协同作用。纤维素对在转化为游离酶时起协同作用，而游离酶主要在野生型状态下起协同作用。发现靶向作用是造成这些特定酶组合观察到的活性升高的主要因素，而邻近效应似乎起着可忽略的作用。

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