Laboratory of Molecular Plant Biology and KU Leuven Plant Institute, KU Leuven, Kasteelpark Arenberg 31, 3001 Leuven, Belgium.
Laboratory of Molecular Plant Biology and KU Leuven Plant Institute, KU Leuven, Kasteelpark Arenberg 31, 3001 Leuven, Belgium.
Carbohydr Polym. 2024 Mar 15;328:121690. doi: 10.1016/j.carbpol.2023.121690. Epub 2023 Dec 15.
World-wide, pathogenic fungi such as Botrytis cinerea cause tremendous yield losses in terms of food production and post-harvest food decay. Many fungi produce inulin-type oligosaccharides (IOSs) from inulin through endo-inulinases which typically show a two domain structure. B.cinerea lacks a two domain endo-inulinase but contains a three domain structure instead. Genome mining revealed three and four domain (d4) enzymes in the fungal kingdom. Here, three and two domain enzymes were compared in their capacity to produce IOSs from inulin. Hill kinetics were observed in three domain enzymes as compared to Michaelis-Menten kinetics in two domain enzymes, suggesting that the N-terminal extension functions as a carbohydrate binding module. Analysis of the IOS product profiles generated from purified GF6, GF12, GF16 and GF18 inulins and extensive sugar docking approaches led to enhanced insights in the active site functioning, revealing subtle differences between the endo-inulinases from Aspergillus niger and B. cinerea. Improved insights in structure-function relationships in fungal endo-inulinases offer opportunities to develop superior enzymes for the production of specific IOS formulations to improve plant and animal health (priming agents, prebiotics).
在全球范围内，病原真菌如 Botrytis cinerea 会导致粮食生产和收获后食品腐烂方面的巨大产量损失。许多真菌通过内切菊粉酶从菊粉中产生菊糖型低聚糖（IOSs），这些酶通常具有两个结构域。然而，B. cinerea 缺乏双结构域内切菊粉酶，而是含有三结构域。基因组挖掘在真菌王国中发现了三结构域和四结构域（d4）酶。在这里，比较了三结构域酶和二结构域酶从菊粉生产 IOSs 的能力。与二结构域酶的米氏动力学相比，三结构域酶表现出 Hill 动力学，这表明 N 端延伸作为碳水化合物结合模块发挥作用。对从纯化的 GF6、GF12、GF16 和 GF18 菊粉中产生的 IOS 产物谱进行分析，并采用广泛的糖对接方法，深入了解了活性位点的功能，揭示了黑曲霉和 B. cinerea 内切菊粉酶之间的细微差异。对真菌内切菊粉酶的结构-功能关系的深入了解为开发生产特定 IOS 配方的优异酶提供了机会，以改善植物和动物健康（启动剂、益生元）。
