Institute for Biology and Biotechnology of Plants, University of Münster, Schlossplatz 8, 48143 Münster, Germany.
ARTES Biotechnology GmbH, Elisabeth-Selbert-Str. 9, 40764 Langenfeld, Germany.
Carbohydr Polym. 2018 Mar 1;183:1-10. doi: 10.1016/j.carbpol.2017.11.015. Epub 2017 Nov 3.
Chitosan is a structurally diverse biopolymer that is commercially derived from chitin by chemical processing, but chitin deacetylases (CDAs) potentially offer a sustainable and more controllable approach allowing the production of chitosans with tailored structures and biological activities. We investigated the CDA from Podospora anserina (PaCDA) which is closely related to Colletotrichum lindemuthianum CDA in the catalytic domain, but unique in having two chitin-binding domains. We produced recombinant PaCDA in Hansenula polymorpha for biochemical characterization and found that the catalytic domain of PaCDA is also functionally similar to C. lindemuthianum CDA, though differing in detail. When studying the enzyme's mode of action on chitin oligomers by quantitative mass-spectrometric sequencing, we found almost all possible sequences up to full deacetylation but with a clear preference for specific products. Deletion muteins lacking one or both CBDs confirmed their proposed function in supporting the enzymatic conversion of the insoluble substrate colloidal chitin.
壳聚糖是一种结构多样的生物聚合物，可通过化学处理从甲壳素商业提取得到，但壳二糖脱乙酰酶（CDAs）可能提供了一种可持续且更可控的方法，可用于生产具有定制结构和生物活性的壳聚糖。我们研究了来自 Podospora anserina 的 CDAs（PaCDA），它在催化结构域与 Colletotrichum lindemuthianum 的 CDA 密切相关，但具有两个壳聚糖结合结构域的特点。我们在 Hansenula polymorpha 中生产重组 PaCDA 进行生化特性分析，发现 PaCDA 的催化结构域在功能上也与 C. lindemuthianum CDA 相似，但细节上有所不同。当通过定量质谱测序研究该酶对壳寡糖的作用模式时，我们发现几乎所有可能的序列都达到了完全脱乙酰化，但对特定产物有明显的偏好。缺失突变体缺乏一个或两个 CBD 证实了它们在支持不溶性底物胶体壳聚糖的酶促转化中的作用。
