Conversion of Thermobifida fusca free exoglucanases into cellulosomal components:: Comparative impact on cellulose-degrading activity
Conversion of Thermobifida fusca free exoglucanases into cellulosomal components:: Comparative impact on cellulose-degrading activity
复制标题
DOI：
10.1016/j.jbiotec.2008.05.003
复制
发表时间：
2008-07-31
影响因子：
4.1
通讯作者：
Bayer, Edward A.
中科院分区：
文献类型：
--
作者：
Caspi, Jonathan;Irwin, Diana;Bayer, Edward A.
纤维素体是由某些厌氧细菌产生的多酶复合物，其表现出对植物细胞壁多糖的有效降解。为了了解它们的水解增强水平，我们正在研究将游离纤维素酶系统转化为纤维素酶体系统的影响。为了实现这一目标，我们正在用已建立特异性的纤维素酶体衍生的锚定蛋白模块替换由好氧细菌Thermobifida fusca产生的天然纤维素酶的细胞纤维素结合模块，以允许其并入定义的“设计者纤维素酶体”。在这篇文章中，我们将不同的锚定蛋白连接到由T. fusca外切葡聚糖酶，Cel 6 B和Cel 48 A。所得到的融合蛋白被证明是有效的，并特异性地结合到他们的匹配的cohesins，并在几个不同的纤维素基板上的活动进行了比较。Cel 6 B中缺乏纤维素结合模块对其在结晶基质上的活性产生有害影响。相比之下，与野生型酶相比，锚定蛋白-听觉家族-48外切葡聚糖酶显示出对羧甲基纤维素和对测试的两种结晶底物的水解活性水平增加。两种外切葡聚糖酶对掺入多纤维素酶体的反应的显著差异表明，家族-48纤维素酶比家族-6酶更适合作为设计者多纤维素酶体组分。(C)2008 Elsevier B. V.保留所有权利。
Cellulosomes are multi-enzyme complexes produced by certain anaerobic bacteria that exhibit efficient degradation of plant cell wall polysaccharides. To understand their enhanced levels of hydrolysis, we are investigating the effects of converting a free-cellulase system into a cellulosomal one. To achieve this end, we are replacing the cell ulose-binding module of the native cellulases, produced by the aerobic bacterium Thermobifida fusca, with a cellulosome-derived dockerin module of established specificity, to allow their incorporation into defined "designer cellulosomes". In this communication, we have attached divergent dockerins to the two exoglucanases produced by T. fusca exoglucanase, Cel6B and Cel48A. The resultant fusion proteins were shown to bind efficiently and specifically to their matching cohesins, and their activities on several different cellulose substrates were compared. The lack of a cellulose-binding module in Cel6B had a deleterious effect on its activity on crystalline substrates. In contrast, the dockerin-hearing family-48 exoglucanase showed increased levels of hydrolytic activity on carboxymethyl cellulose and on both Crystalline Substrates tested, compared to the wild-type enzyme. The marked difference in the response of the two exoglucanases to incorporation into a cellulosome, Suggests that the family-48 cellulase is more appropriate than the family-6 enzyme as a designer cellulosome component. (C) 2008 Elsevier B.V. All rights reserved.