QM/MM Studies into the H2O2-Dependent Activity of Lytic Polysaccharide Monooxygenases: Evidence for the Formation of a Caged Hydroxyl Radical Intermediate

Lytic polysaccharide monooxygenases (LPMOs) are promising enzymes for the conversion of lignocellulosic biomass into biofuels and biomaterials. Classically considered oxygenases, recent work suggests that H2O2 can, under certain circumstances, also be a potential substrate. Here we present a detailed mechanism of the activation of H2O2 by a C4-acting LPMO using small-model DFT and QM/MM calculations. We show that there is an efficient mechanism to break the O-O bond of H2O2 with a low barrier of 5.8 kcal/mol, via a one-electron transfer from the LPMO-Cu(I) site to form an HO center dot radical, stabilized by hydrogen bonding interactions. Our QM/MM calculations further show that the H-bonding machinery of the enzyme directs the HO center dot radical to abstract a hydrogen atom from the Cu(II) OH unit rather from substrate in what is essentially a caged-radical reaction, thereby forming a Cu(II)-oxyl species. The Cu(II)-oxyl species then exclusively oxidizes the C4-H bond due to the suitable position of the substrate. Our calculations also suggest that the C4-hydroxylated intermediate can be efficiently hydrolyzed in water, and this process does not require enzymatic catalysis.