Synergistic Mechanism Of Lignocellulolytic Enzyme Systems On Herbaceous Biomass

Lignocellulose is considered as an abundant renewable resource. Due to its heterogeneity and complexity, the complete degradation of lignocellulose requires the synergistic action of diverse enzymes. In nature, some microorganisms are capable of producing complex lignocellulolytic enzyme systems. An in-depth study of the synergistic action and mechanism of lignocellulolytic enzymes is of great significance to the development of more efficient enzyme preparations and the improvement of biomass conversion.In this study, the enzyme systems of wood-decay basidiomycete Schizophyllum commune SH12 degrading Jerusalem artichoke stalk were investigated. Compared with model white-rot and brown-rot fungi, S. commune displayed considerably higher levels of hydrolytic enzyme activities, including cellulases, hemicellulases and pectinases. While it preferentially degraded cellulose and hemicellulose, S. commune also employed hydroxyl radicals to modify the lignin polymer.S. commune produced a set of peroxide-generating enzymes and iron-reducing enzymes, potentially supporting the generation of hydroxyl radicals. The cellulose converions of S. commune enzymes were 2.5-3.8 times as high as those of commercial enzymes in the hydrolysis of delignified lignocellulosic biomass. LC-MS/MS analysis of the secretomes revealed that S. commune produced a high diversity of carbohydrate-degrading enzymes, especially hemicellulases and pectinases acting on polysaccharide backbones and side chains. In addition, multiple non-hydrolytic proteins were identified, including polysaccharide monooxygenases (PMOs) and expansins. The significant saccharification efficiency of S. commune enzymes could be attributed to the synergism between these proteins.In natural ecosystems, biodegradation of lignocellulosic biomass is usually accomplished by intricate consortia of diverse microorganisms. Metaproteomic approaches were used to study the enzyme systems and synergy of an efficient lignocellulolytic microbial consortium EMSD5 during growth on corn stover. The expression of most degradative enzymes in EMSD5 was substrate-dependent. A wide range of lignocellulolytic enzymes from diverse Firmicutes bacteria were induced by corn stover, and acted in a synergistic way. Quantitative proteome data in this study revealed the sequential secretion of enzymes with differential expression levels. A core subset of degradative enzymes showing significant abundance increase were identified during the deconstruction process of corn stover, including xylanases, cellobiohydrolases and endoglucanases. In addition, some transport proteins and hypothetical proteins were coordinately expressed with glycoside hydrolases, suggesting a role in the lignocellulose degradation by EMSD5. EMSD5 enzymes promoted the hydrolysis of pretreated corn stover by commercial cellulases from Trichoderma reesei, with a degree of synergism of 1.96.Two CBM1-containing PMOs, rPc2980158 and rPc3003776 were heterologously expressed, and their synergy with cellulases was characterized. Both PMOs improved the hydrolytic efficiency of T. reesei cellulase enzymes on microcrystalline cellulose and pretreated switchgrass. Their activities against pure cellulose depended on electron donors, and were potentiated by multiple divalent metal ions. The intact rPc2980158 demonstrated stronger adsorption strength on all substrates than CBM1-deleted variant, indicating the promotion of substrate-binding capacities of PMOs by CBMl. The intact protein improved the cellulose hydrolysis of pure cellulose and pretreated lignocellulosic biomass to much greater extents than CBM1-deleted variant. The improvements in cellulose hydrolysis by addition of rPc2980158 correlated well with its adsorption capacity. These results demonstrated that the specific adsorption of CBM1 to cellulose played a major role in the binding of PMOs to lignocellulosic substrates and boosted the synergy between PMOs and cellulases.A potential carbohydrate-active enzyme Sc2146679 was identified in the secretome of S. commune. Sc2146679 was heterologously expressed and its substrate specificities were investigated. rSc2146679 acted as a glycoside hydrolase, and showed hydrolytic activities on amylopectin, amylose, cellulose and laminarin. rSc2146679 mainly acted on α-1,4-glycosidic bonds, but also cleaved β-1,4- and β-1,3-glycosidic bonds. rSc2146679 converted 19% of amylopectin to glucose in 72 hours.