The “life-span” of lytic polysaccharide monooxygenases (LPMOs) correlates to the number of turnovers in the reductant peroxidase reaction

Lytic polysaccharide monooxygenases (LPMOs) are monocopper enzymes that degrade the insoluble crystalline polysaccharides cellulose and chitin. Besides the H₂O₂ cosubstrate, the cleavage of glycosidic bonds by LPMOs depends on the presence of a reductant needed to bring the enzyme into its reduced, catalytically active Cu(I) state. Reduced LPMOs that are not bound to substrate catalyze reductant peroxidase reactions, which may lead to oxidative damage and irreversible inactivation of the enzyme. However, the kinetics of this reaction remain largely unknown, as do possible variations between LPMOs belonging to different families. Here, we describe the kinetic characterization of two fungal family AA9 LPMOs, TrAA9A of Trichoderma reesei and NcAA9C of Neurospora crassa, and two bacterial AA10 LPMOs, ScAA10C of Streptomyces coelicolor and SmAA10A of Serratia marcescens. We found peroxidation of ascorbic acid and methyl-hydroquinone resulted in the same probability of LPMO inactivation (p_i), suggesting that inactivation is independent of the nature of the reductant. We showed the fungal enzymes were clearly more resistant toward inactivation, having p_i values of less than 0.01, whereas the p_i for SmAA10A was an order of magnitude higher. However, the fungal enzymes also showed higher catalytic efficiencies (k_cat/K_M(H2O2)) for the reductant peroxidase reaction. This inverse linear correlation between the k_cat/K_M(H2O2) and p_i suggests that, although having different life spans in terms of the number of turnovers in the reductant peroxidase reaction, LPMOs that are not bound to substrates have similar half-lives. These findings have not only potential biological but also industrial implications.

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