UR1268 BIA, INRAE, F-44316 Nantes, France.
INRAE, Aix Marseille Univ., UMR BBF, F-13009 Marseille, France.
Biomacromolecules. 2023 Jul 10;24(7):3246-3255. doi: 10.1021/acs.biomac.3c00303. Epub 2023 Jun 16.
Lytic polysaccharide monooxygenase (LPMO) enzymes have recently shaken up our knowledge of the enzymatic degradation of biopolymers and cellulose in particular. This unique class of metalloenzymes cleaves cellulose and other recalcitrant polysaccharides using an oxidative mechanism. Despite their potential in biomass saccharification and cellulose fibrillation, the detailed mode of action of LPMOs at the surface of cellulose fibers still remains poorly understood and highly challenging to investigate. In this study, we first determined the optimal parameters (temperature, pH, enzyme concentration, and pulp consistency) of LPMO action on the cellulose fibers by analyzing the changes in molar mass distribution of solubilized fibers using high performance size exclusion chromatography (HPSEC). Using an experimental design approach with a fungal LPMO from the AA9 family (LPMO9H) and cotton fibers, we revealed a maximum decrease in molar mass at 26.6 °C and pH 5.5, with 1.6% enzyme loading in dilute cellulose dispersions (100 mg of cellulose at 0.5% ). These optimal conditions were used to further investigate the effect of LPMO9H on the cellulosic fiber structure. Direct visualization of the fiber surface by scanning electron microscopy (SEM) revealed that LPMO9H created cracks on the cellulose surface while it attacked tension regions that triggered the rearrangement of cellulose chains. Solid-state NMR indicated that LPMO9H increased the lateral fibril dimension and created novel accessible surfaces. This study confirms the LPMO-driven disruption of cellulose fibers and extends our knowledge of the mechanism underlying such modifications. We hypothesize that the oxidative cleavage at the surface of the fibers releases the tension stress with loosening of the fiber structure and peeling of the surface, thereby increasing the accessibility and facilitating fibrillation.
溶细胞聚糖单加氧酶（LPMO）酶最近颠覆了我们对生物聚合物，尤其是纤维素的酶促降解的认识。这种独特的金属酶类使用氧化机制切割纤维素和其他难降解的多糖。尽管它们在生物质糖化和纤维素原纤化方面具有潜力，但 LPMO 在纤维素纤维表面的详细作用机制仍然知之甚少，并且极具挑战性。在这项研究中，我们首先通过使用高效尺寸排阻色谱（HPSEC）分析溶解纤维的摩尔质量分布，确定了 LPMO 在纤维素纤维上作用的最佳参数（温度、pH 值、酶浓度和纸浆浓度）。通过使用真菌 AA9 家族的 LPMO（LPMO9H）和棉纤维的实验设计方法，我们揭示了在 26.6°C 和 pH5.5 下摩尔质量最大降低，在稀纤维素分散体（纤维素 100mg 于 0.5%）中酶负载为 1.6%。这些最佳条件用于进一步研究 LPMO9H 对纤维素纤维结构的影响。通过扫描电子显微镜（SEM）直接观察纤维表面，发现 LPMO9H 在攻击引发纤维素链重排的张力区域的同时，在纤维素表面产生了裂纹。固态 NMR 表明 LPMO9H 增加了侧向原纤尺寸并创造了新的可及表面。这项研究证实了 LPMO 驱动的纤维素纤维的破坏，并扩展了我们对这种修饰背后的机制的认识。我们假设纤维表面的氧化切割释放了张力，从而使纤维结构变松并剥落表面，从而增加了可及性并促进了原纤化。
