A biochemical comparison of fungal GH6 cellobiohydrolases
A biochemical comparison of fungal GH6 cellobiohydrolases
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DOI：
10.1042/bcj20190185
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发表时间：
2019-08-15
影响因子：
4.1
通讯作者：
Westh, Peter
中科院分区：
文献类型：
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作者：
Christensen, Stefan Jarl;Krogh, Kristian Bertel Romer Morkeberg;Westh, Peter
来自糖苷水解酶家族6（GH6）的纤维二糖水解酶（CBH）是许多纤维素分解真菌分泌组的重要组成部分。它们也具有技术意义，特别是因为它们是用于木质纤维素生物质的工业分解的酶混合物的一部分。然而，GH6 CBH的功能研究是稀缺的，集中在几个模型酶。为了阐明GH6 CBH之间的功能广度，我们进行了比较生化研究的七个GH6 CBH来自真菌生活在不同的栖息地，除了一个酶的变体。通过系统发育分析对酶序列进行了研究，以确保它们在遗传学上不密切相关。所选的酶都异源表达在Aspergilluscaureus，纯化和彻底的生化特性。这种方法允许功能数据的直接比较，结果显示了很大的差异。例如，对纤维素的吸附能力跨越两个数量级和动力学参数，来自两个独立的稳态方法也有显着差异。虽然不同的功能参数涵盖范围很广，但它们并不是独立的，因为它们在两极之间平行变化。其中一个极点的特征是强基质相互作用，高吸附容量和低周转数，而另一个极点的特征是弱基质相互作用，吸附差，高周转。所研究的酶基本上定义了这两个对立之间的连续体，这种功能参数的缩放提出了有趣的问题，关于功能可塑性和进化的GH6 CBH。
Cellobiohydrolases (CBHs) from glycoside hydrolase family 6 (GH6) make up an important part of the secretome in many cellulolytic fungi. They are also of technical interest, particularly because they are part of the enzyme cocktails that are used for the industrial breakdown of lignocellulosic biomass. Nevertheless, functional studies of GH6 CBHs are scarce and focused on a few model enzymes. To elucidate functional breadth among GH6 CBHs, we conducted a comparative biochemical study of seven GH6 CBHs originating from fungi living in different habitats, in addition to one enzyme variant. The enzyme sequences were investigated by phylogenetic analyses to ensure that they were not closely related phylogenetically. The selected enzymes were all heterologously expressed in Aspergillus oryzae, purified and thoroughly characterized biochemically. This approach allowed direct comparisons of functional data, and the results revealed substantial variability. For example, the adsorption capacity on cellulose spanned two orders of magnitude and kinetic parameters, derived from two independent steady-state methods also varied significantly. While the different functional parameters covered wide ranges, they were not independent since they changed in parallel between two poles. One pole was characterized by strong substrate interactions, high adsorption capacity and low turnover number while the other showed weak substrate interactions, poor adsorption and high turnover. The investigated enzymes essentially defined a continuum between these two opposites, and this scaling of functional parameters raises interesting questions regarding functional plasticity and evolution of GH6 CBHs.