TY - JOUR

T1 - AA16 Oxidoreductases Boost Cellulose-Active AA9 Lytic Polysaccharide Monooxygenases from Myceliophthora thermophila

AU - Sun, Peicheng

AU - Huang, Zhiyu

AU - Banerjee, Sanchari

AU - Kadowaki, Marco A. S.

AU - Veersma, Romy J.

AU - Magri, Silvia

AU - Hilgers, Roelant

AU - Muderspach, Sebastian J.

AU - Laurent, Christophe V. F. P.

AU - Ludwig, Roland

AU - Cannella, David

AU - Lo Leggio, Leila

AU - van Berkel, Willem J. H.

AU - Kabel, Mirjam A.

PY - 2023

Y1 - 2023

N2 - Copper-dependent lytic polysaccharide monooxygenases (LPMOs) classified in Auxiliary Activity (AA) families are considered indispensable as synergistic partners for cellulolytic enzymes to saccharify recalcitrant lignocellulosic plant biomass. In this study, we characterized two fungal oxidoreductases from the new AA16 family. We found that MtAA16A from Myceliophthora thermophila and AnAA16A from Aspergillus nidulans did not catalyze the oxidative cleavage of oligo-and polysaccharides. Indeed, the MtAA16A crystal structure showed a fairly LPMO-typical histidine brace active site, but the cellulose-acting LPMO-typical flat aromatic surface parallel to the histidine brace region was lacking. Further, we showed that both AA16 proteins are able to oxidize low-molecular-weight reductants to produce H2O2. The oxidase activity of the AA16s substantially boosted cellulose degradation by four AA9 LPMOs from M. thermophila (MtLPMO9s) but not by three AA9 LPMOs from Neurospora crassa (NcLPMO9s). The interplay with MtLPMO9s is explained by the H2O2-producing capability of the AA16s, which, in the presence of cellulose, allows the MtLPMO9s to optimally drive their peroxygenase activity. Replacement of MtAA16A by glucose oxidase (AnGOX) with the same H2O2-producing activity could only achieve less than 50% of the boosting effect achieved by MtAA16A, and earlier MtLPMO9B inactivation (6 h) was observed. To explain these results, we hypothesized that the delivery of AA16produced H2O2 to the MtLPMO9s is facilitated by protein-protein interaction. Our findings provide new insights into the functions of copper-dependent enzymes and contribute to a further understanding of the interplay of oxidative enzymes within fungal systems to degrade lignocellulose.

AB - Copper-dependent lytic polysaccharide monooxygenases (LPMOs) classified in Auxiliary Activity (AA) families are considered indispensable as synergistic partners for cellulolytic enzymes to saccharify recalcitrant lignocellulosic plant biomass. In this study, we characterized two fungal oxidoreductases from the new AA16 family. We found that MtAA16A from Myceliophthora thermophila and AnAA16A from Aspergillus nidulans did not catalyze the oxidative cleavage of oligo-and polysaccharides. Indeed, the MtAA16A crystal structure showed a fairly LPMO-typical histidine brace active site, but the cellulose-acting LPMO-typical flat aromatic surface parallel to the histidine brace region was lacking. Further, we showed that both AA16 proteins are able to oxidize low-molecular-weight reductants to produce H2O2. The oxidase activity of the AA16s substantially boosted cellulose degradation by four AA9 LPMOs from M. thermophila (MtLPMO9s) but not by three AA9 LPMOs from Neurospora crassa (NcLPMO9s). The interplay with MtLPMO9s is explained by the H2O2-producing capability of the AA16s, which, in the presence of cellulose, allows the MtLPMO9s to optimally drive their peroxygenase activity. Replacement of MtAA16A by glucose oxidase (AnGOX) with the same H2O2-producing activity could only achieve less than 50% of the boosting effect achieved by MtAA16A, and earlier MtLPMO9B inactivation (6 h) was observed. To explain these results, we hypothesized that the delivery of AA16produced H2O2 to the MtLPMO9s is facilitated by protein-protein interaction. Our findings provide new insights into the functions of copper-dependent enzymes and contribute to a further understanding of the interplay of oxidative enzymes within fungal systems to degrade lignocellulose.

KW - cellulose

KW - Carbohydrate-Active enZyme

KW - copper-dependent oxidoreductase

KW - fungal auxiliary activity family

KW - hydrogen peroxide

KW - lytic polysaccharide monooxygenase

KW - protein structure

KW - CELLOBIOSE DEHYDROGENASE

KW - OXIDATIVE CLEAVAGE

KW - WEB SERVER

KW - ENZYMES

KW - DEGRADATION

KW - BIOMASS

KW - DISCOVERY

KW - OXIDASE

KW - DALI

U2 - 10.1021/acscatal.3c00874

DO - 10.1021/acscatal.3c00874

M3 - Journal article

C2 - 37066045

VL - 13

SP - 4454

EP - 4467

JO - ACS Catalysis

JF - ACS Catalysis

SN - 2155-5435

IS - 7

ER -