Polymer ultrastructure governs AA9 lytic polysaccharide monooxygenases functionalization and deconstruction efficacy on cellulose nano-crystals

[Display omitted] •AA9 LPMO-mediated cracking and functionalization of cellulose nanocrystals.•Cellulose ultrastructure affects AA9 LPMOs regioselectivity and catalysis.•AA9 LPMOs overcome the high dry matter effect.•Differential preference of amorphous and crystalline cellulose among AA9 LPMOs. Lytic Polysaccharide MonoOxygenases display great variability towards cellulose ultrastructure while performing oxidative functionalization of the polymers. Aiming at employing AA9-LPMOs for isolation of cellulose nano-crystals (CNCs), the ratio between functionalization/crystalline degradation became a crucial parameter. Here are reported the constraints posed by the substrate ultrastructure on the activity of seven different AA9 LPMOs representative of various regioselectivity and substrate affinity: TtAA9E, TaAA9A, PcAA9D, MtAA9A, MtAA9D, MtAA9I-CBM and MtAA9J. The substrate crystallinity and dry matter loading greatly affected the seven AA9s in an enzyme-specific manner, impacting their efficiency for CNCs functionalization purposes. X-ray diffraction pattern analyses were used to assess the cracking efficacy of the enzymatic treatment to de-crystallize CNCs, revealing that those AA9s with minor efficiency in releasing oligosaccharides resulted instead the most disruptive towards the crystal lattice and in reducing the particle sizes. These non-catalytic effects were found comparable with the one caused by the expansin BsEXLX1 enzyme.