Oxidative cleavage of polysaccharides by monocopper enzymes depends on H2O2

Enzymes currently known as lytic polysaccharide monooxygenases (LPMOs) play an important role in the conversion of recalcitrant polysaccharides, but their mode of action has remained largely enigmatic. It is generally believed that catalysis by LPMOs requires molecular oxygen and a reductant that delivers two electrons per catalytic cycle. Using enzyme assays, mass spectrometry and experiments with labeled oxygen atoms, we show here that H 2 O 2 , rather than O 2 , is the preferred co-substrate of LPMOs. By controlling H 2 O 2 supply, stable reaction kinetics are achieved, the LPMOs work in the absence of O 2 , and the reductant is consumed in priming rather than in stoichiometric amounts. The use of H 2 O 2 by a monocopper enzyme that is otherwise cofactor-free offers new perspectives regarding the mode of action of copper enzymes. Furthermore, these findings have implications for the enzymatic conversion of biomass in Nature and in industrial biorefining.