The Molecular Mechanisms of Oxygen Activation and Hydrogen Peroxide Formation in Lytic Polysaccharide Monooxygenases

TY - JOUR

T1 - The Molecular Mechanisms of Oxygen Activation and Hydrogen Peroxide Formation in Lytic Polysaccharide Monooxygenases

AU - Wang, Binju

AU - Walton, Paul Howard

AU - Rovira, Carme

N1 - © 2019 American Chemical Society. This is an author-produced version of the published paper. Uploaded in accordance with the publisher’s self-archiving policy. Further copying may not be permitted; contact the publisher for details.

PY - 2019/4/22

Y1 - 2019/4/22

N2 - Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes for the degradation of recalcitrant polysaccharides such as chitin and cellulose. Unlike classical hydrolytic enzymes (cellulases), LPMOs catalyze the cleavage of the glycosidic bond via an oxidative mechanism using oxygen and a reductant. The full enzymatic molecular mechanisms, starting from the initial electron transfer from a reductant to oxygen activation and hydrogen peroxide formation, are not yet understood. Using QM/MM metadynamics simulations, we have uncovered the oxygen activation mechanisms by LPMO in the presence of ascorbic acid, one of the most-used reductants in LPMOs assays. Our simulations capture the sequential formation of Cu(II)-O2- and Cu(II)-OOH- intermediates via facile H-atom abstraction from ascorbate. By investigating all the possible reaction pathways from the Cu(II)−OOH- intermediate, we ruled out Cu(II)-O•- formation via direct O-O cleavage of Cu(II)-OOH-. Meanwhile, we identified a possible pathway in which the proximal oxygen atom of Cu(II)−OOH- abstracts a hydrogen atom from ascorbate, leading to Cu(I) and H2O2. The “in situ” generated H2O2 either converts to LPMO-Cu(II)-O•- via a homolytic reaction, or diffuses into the bulk water in an uncoupled pathway. The competition of these two pathways is strongly dependent on the binding of the carbohydrate substrate, which plays a role in barricading the “in situ” generated H2O2 molecule, preventing its diffusion from the active site into the bulk water. Based on the present results, we propose a catalytic cycle of LPMOs that is consistent with the experimental information available. In particular, it explains the enigmatic substrate-dependence of the reactivity of the LPMO with H2O2.

AB - Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes for the degradation of recalcitrant polysaccharides such as chitin and cellulose. Unlike classical hydrolytic enzymes (cellulases), LPMOs catalyze the cleavage of the glycosidic bond via an oxidative mechanism using oxygen and a reductant. The full enzymatic molecular mechanisms, starting from the initial electron transfer from a reductant to oxygen activation and hydrogen peroxide formation, are not yet understood. Using QM/MM metadynamics simulations, we have uncovered the oxygen activation mechanisms by LPMO in the presence of ascorbic acid, one of the most-used reductants in LPMOs assays. Our simulations capture the sequential formation of Cu(II)-O2- and Cu(II)-OOH- intermediates via facile H-atom abstraction from ascorbate. By investigating all the possible reaction pathways from the Cu(II)−OOH- intermediate, we ruled out Cu(II)-O•- formation via direct O-O cleavage of Cu(II)-OOH-. Meanwhile, we identified a possible pathway in which the proximal oxygen atom of Cu(II)−OOH- abstracts a hydrogen atom from ascorbate, leading to Cu(I) and H2O2. The “in situ” generated H2O2 either converts to LPMO-Cu(II)-O•- via a homolytic reaction, or diffuses into the bulk water in an uncoupled pathway. The competition of these two pathways is strongly dependent on the binding of the carbohydrate substrate, which plays a role in barricading the “in situ” generated H2O2 molecule, preventing its diffusion from the active site into the bulk water. Based on the present results, we propose a catalytic cycle of LPMOs that is consistent with the experimental information available. In particular, it explains the enigmatic substrate-dependence of the reactivity of the LPMO with H2O2.

U2 - 10.1021/acscatal.9b00778

DO - 10.1021/acscatal.9b00778

M3 - Article

SN - 2155-5435

JO - ACS Catalysis

JF - ACS Catalysis

ER -