Abstract

Arabinanases from glycoside hydrolase family GH93 are enzymes with exo-activity that hydrolyze the α-1,5 bonds between arabinose residues present on arabinan. Currently, several initiatives aiming to use byproducts rich in arabinan such as pectin and sugar beet pulp as raw material to produce various compounds of interest are being developed. However, it is necessary to use robust enzymes that have an optimal performance under pH and temperature conditions used in the industrial processes. In this work, the first GH93 from the thermophilic fungus Thermothielavioides terrestris (Abn93T) was heterologously expressed in Aspergillus nidulans, purified and biochemically characterized. The enzyme is a thermophilic glycoprotein (optimum activity at 70 °C) with prolonged stability in acid pHs (4.0 to 6.5). The presence of glycosylation affected slightly the hydrolytic capacity of the enzyme, which was further increased by 34% in the presence of 1 mM CoCl₂. Small-angle X-ray scattering results show that Abn93T is a globular-like-shaped protein with a slight bulge at one end. The hydrolytic mechanism of the enzyme was elucidated using capillary zone electrophoresis and molecular docking calculations. Abn93T has an ability to produce (in synergism with arabinofuranosidases) arabinose and arabinobiose from sugar beet arabinan, which can be explored as fermentable sugars and prebiotics.

Key points

• Thermophilic exo-arabinanase from family GH93

• Molecular basis of arabinan depolymerization

中文翻译：

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一种新型的嗜热拟南芥嗜热外阿拉伯糖苷酶的功能表征。

摘要

糖苷水解酶家族GH93的阿拉伯聚糖酶是具有exo活性的酶，可水解阿拉伯聚糖上存在的阿拉伯糖残基之间的α-1,5键。当前，正在开发一些旨在使用富含阿拉伯聚糖的副产物如果胶和甜菜浆作为原料来生产各种目的化合物的举措。但是，必须使用在工业过程中使用的pH和温度条件下具有最佳性能的健壮酶。在这项工作中，嗜热性真菌Therththielavioides terrestris（Abn93T）的第一个GH93在构巢曲霉中异源表达。，纯化和生化特性。该酶是嗜热的糖蛋白（在70°C时具有最佳活性），在酸性pH值（4.0至6.5）下具有较长的稳定性。糖基化的存在对酶的水解能力稍有影响，在1 mM CoCl ₂的存在下，其水解能力进一步提高了34％。小角X射线散射结果表明，Abn93T是一种球状蛋白，一端稍有凸起。使用毛细管区带电泳和分子对接计算阐明了酶的水解机理。Abn93T具有从甜菜阿拉伯糖产生阿拉伯糖和阿拉伯二糖（与阿拉伯呋喃糖苷酶协同作用）的能力，可以将其开发为可发酵糖和益生元。

关键点

• GH93家族的嗜热外阿拉伯聚糖酶

•阿拉伯聚糖解聚的分子基础