Department of Biology and Biological Engineering, Division of Industrial Biotechnology, Chalmers University of Technology, Gothenburg, Sweden
Wallenberg Wood Science Center, Chalmers University of Technology, Gothenburg, Sweden.
Appl Environ Microbiol. 2019 Nov 14;85(23). doi: 10.1128/AEM.01408-19. Print 2019 Dec 1.
The thermophilic biomass-degrader exhibits poor growth on cellulose but excellent growth on hemicelluloses as the sole carbon source. This is surprising considering that its genome encodes eight lytic polysaccharide monooxygenases (LPMOs) from auxiliary activity family 9 (AA9), enzymes known for their high potential in accelerating cellulose depolymerization. We characterized four of the eight ( AA9s) AA9s, namely, AA9A, AA9B, AA9F, and AA9H, to gain a deeper understanding about their roles in the fungus. The characterized AA9s were active on hemicelluloses, including xylan, glucomannan, and xyloglucan, and furthermore, in accordance with transcriptomics data, differed in substrate specificity. Of the AA9s, AA9H is unique, as it preferentially cleaves residual xylan in phosphoric acid-swollen cellulose (PASC). Moreover, when exposed to cellulose-xylan blends, AA9H shows a preference for xylan and for releasing (oxidized) xylooligosaccharides. The cellulose dependence of the xylan activity suggests that a flat conformation, with rigidity similar to that of cellulose microfibrils, is a prerequisite for productive interaction between xylan and the catalytic surface of the LPMO. AA9H showed a similar trend on xyloglucan, underpinning the suggestion that LPMO activity on hemicelluloses strongly depends on the polymers' physicochemical context and conformation. Our results support the notion that LPMO multiplicity in fungal genomes relates to the large variety of copolymeric polysaccharide arrangements occurring in the plant cell wall. The LPMOs (AA9s) showed activity on a broad range of soluble and insoluble substrates, suggesting their involvement in various steps of biomass degradation besides cellulose decomposition. Our results indicate that the fungal AA9 family is more diverse than originally thought and able to degrade almost any kind of plant cell wall polysaccharide. The discovery of an AA9 that preferentially cleaves xylan enhances our understanding of the physiological roles of LPMOs and enables the use of xylan-specific LPMOs in future applications.
嗜热生物质降解菌在纤维素上的生长较差，但在半纤维素上的生长极好，因为半纤维素是其唯一的碳源。考虑到其基因组编码了 8 种来自辅助活性家族 9（AA9）的溶菌多糖单加氧酶（LPMO），这些酶以在加速纤维素解聚方面的高潜力而闻名，这种情况令人惊讶。我们对这 8 种 LPMO 中的 4 种（AA9s），即 AA9A、AA9B、AA9F 和 AA9H 进行了表征，以更深入地了解它们在真菌中的作用。这 4 种 AA9s 在半纤维素（包括木聚糖、葡甘露聚糖和木葡聚糖）上具有活性，此外，根据转录组学数据，它们在底物特异性上有所不同。在这 4 种 AA9s 中，AA9H 是独特的，因为它优先切割磷酸化膨胀纤维素（PASC）中的残留木聚糖。此外，当暴露在纤维素-木聚糖混合物中时，AA9H 优先作用于木聚糖并释放（氧化）木寡糖。木聚糖活性对纤维素的依赖性表明，一种具有类似于纤维素微纤丝的刚性的平面构象，是木聚糖与 LPMO 催化表面之间进行生产性相互作用的前提条件。AA9H 在木葡聚糖上也表现出类似的趋势，这支持了这样一种观点，即 LPMO 在半纤维素上的活性强烈依赖于聚合物的物理化学环境和构象。我们的结果支持这样一种观点，即真菌基因组中 LPMO 的多样性与植物细胞壁中存在的各种共聚多糖排列有关。LPMO（AA9s）在广泛的可溶性和不溶性底物上表现出活性，这表明它们除了参与纤维素分解外，还参与了生物质降解的各个步骤。我们的结果表明，真菌 AA9 家族比最初想象的更加多样化，能够降解几乎任何类型的植物细胞壁多糖。发现一种优先切割木聚糖的 AA9，增强了我们对 LPMO 生理作用的理解，并使木聚糖特异性 LPMO 能够在未来的应用中使用。
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