Optimized Lytic Polysaccharide Monooxygenase Action Increases Fiber Accessibility and Fibrillation by Releasing Tension Stress in Cellulose Cotton Fibers

Lyticpolysaccharide monooxygenase (LPMO) enzymes have recentlyshaken up our knowledge of the enzymatic degradation of biopolymersand cellulose in particular. This unique class of metalloenzymes cleavescellulose and other recalcitrant polysaccharides using an oxidativemechanism. Despite their potential in biomass saccharification andcellulose fibrillation, the detailed mode of action of LPMOs at thesurface of cellulose fibers still remains poorly understood and highlychallenging to investigate. In this study, we first determined theoptimal parameters (temperature, pH, enzyme concentration, and pulpconsistency) of LPMO action on the cellulose fibers by analyzing thechanges in molar mass distribution of solubilized fibers using highperformance size exclusion chromatography (HPSEC). Using an experimentaldesign approach with a fungal LPMO from the AA9 family (PaLPMO9H) and cotton fibers, we revealed a maximum decrease in molarmass at 26.6 degrees C and pH 5.5, with 1.6% w/w enzymeloading in dilute cellulose dispersions (100 mg of cellulose at 0.5% w/v). These optimal conditions were used to further investigatethe effect of PaLPMO9H on the cellulosic fiber structure.Direct visualization of the fiber surface by scanning electron microscopy(SEM) revealed that PaLPMO9H created cracks on thecellulose surface while it attacked tension regions that triggeredthe rearrangement of cellulose chains. Solid-state NMR indicated that PaLPMO9H increased the lateral fibril dimension and creatednovel accessible surfaces. This study confirms the LPMO-driven disruptionof cellulose fibers and extends our knowledge of the mechanism underlyingsuch modifications. We hypothesize that the oxidative cleavage atthe surface of the fibers releases the tension stress with looseningof the fiber structure and peeling of the surface, thereby increasingthe accessibility and facilitating fibrillation.

Automated summary

This study determined the optimal conditions for LPMO action on cellulose fibers and investigated the effect of PaLPMO9H on the fiber structure. The results showed that PaLPMO9H created cracks on the cellulose surface and increased the lateral fibril dimension. This study confirms the disruptive role of LPMO in cellulose fibers and extends our understanding of the underlying mechanisms.