Background The ascomycete fungus Podospora anserina has been appreciated for its targeted carbohydrate-active enzymatic arsenal. As a late colonizer of herbivorous dung, the fungus acts specifically on the more recalcitrant fraction of lignocellulose and this lignin-rich biotope might have resulted in the evolution of ligninolytic activities. However, the lignin-degrading abilities of the fungus have not been demonstrated by chemical analyses at the molecular level and are, thus far, solely based on genome and secretome predictions. To evaluate whether P. anserina might provide a novel source of lignin-active enzymes to tap into for potential biotechnological applications, we comprehensively mapped wheat straw lignin during fungal growth and characterized the fungal secretome. Results Quantitative 13C lignin internal standard py-GC-MS analysis showed substantial lignin removal during the 7 days of fungal growth (24% w/w), though carbohydrates were preferably targeted (58% w/w removal). Structural characterization of residual lignin by using py-GC-MS and HSQC NMR analyses demonstrated that Cα-oxidized substructures significantly increased through fungal action, while intact β-O-4' aryl ether linkages, p-coumarate and ferulate moieties decreased, albeit to lesser extents than observed for the action of basidiomycetes. Proteomic analysis indicated that the presence of lignin induced considerable changes in the secretome of P. anserina. This was particularly reflected in a strong reduction of cellulases and galactomannanases, while H2O2-producing enzymes clearly increased. The latter enzymes, together with laccases, were likely involved in the observed ligninolysis. Conclusions For the first time, we provide unambiguous evidence for the ligninolytic activity of the ascomycete fungus P. anserina and expand the view on its enzymatic repertoire beyond carbohydrate degradation. Our results can be of significance for the development of biological lignin conversion technologies by contributing to the quest for novel lignin-active enzymes and organisms.

中文翻译：

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子囊菌 Podospora anserina 木质素分解活性的证据。


背景 子囊菌 Podospora anserina 因其具有靶向碳水化合物活性的酶库而受到赞赏。作为草食性粪便的晚期定植者，真菌专门作用于木质纤维素中更顽固的部分，这种富含木质素的生境可能导致了木质素分解活性的进化。然而，真菌的木质素降解能力尚未通过分子水平的化学分析得到证实，并且迄今为止仅基于基因组和分泌组预测。为了评估 P. anserina 是否可能提供一种新的木质素活性酶来源以用于潜在的生物技术应用，我们全面绘制了真菌生长过程中的麦秆木质素图谱并表征了​​真菌分泌组。结果 定量 13C 木质素内标 py-GC-MS 分析显示，在真菌生长的 7 天期间，木质素被大量去除（24% w/w），但优选以碳水化合物为目标（58% w/w 去除）。使用 py-GC-MS 和 HSQC NMR 分析对残留木质素进行结构表征表明，Cα 氧化的亚结构通过真菌作用显着增加，而完整的 β-O-4' 芳基醚键、对香豆酸和阿魏酸部分则减少，尽管比担子菌的作用观察到的程度要小。蛋白质组学分析表明，木质素的存在引起鹅绒毛蚁分泌组的显着变化。这尤其体现在纤维素酶和半乳甘露聚糖酶的大幅减少，而产生 H2O2 的酶明显增加。后一种酶与漆酶一起可能参与了观察到的木质素分解。 结论 我们首次为子囊菌 P. anserina 的木质素分解活性提供了明确的证据，并扩展了对其酶谱范围超越碳水化合物降解的认识。我们的研究结果有助于寻找新型木质素活性酶和生物体，对生物木质素转化技术的发展具有重要意义。