Comparison of C4-oxidizing and C1/C4-oxidizing AA9 LPMOs in substrate adsorption, H2O2-driven activity and synergy with cellulase on celluloses of different crystallinity.

Two C1/C4-oxidizing AA9 lytic polysaccharide monooxygenases (AA9 LPMOs), AoLPMO9A and AoLPMO9B, and one C4-oxidizing AoLPMO9C from Aspergillus oryzae, were characterized and compared with the well-studied C4-oxidizing NcLPMO9C. NcLPMO9C and AoLPMO9C harboring carbohydrate-binding module 1 (CBM1) exhibited much stronger adsorption capacity than AoLPMO9A and B without CBM1. The binding affinity is crucial for the efficacy of H2O2 as cosubstrate and oxidative activity of AA9 LPMOs on crystalline cellulose. C4-oxidizing AA9 LPMOs had a striking boosting effect on cellobiohydrolase I (CBHI), while C1/C4-oxidizing AA9 LPMOs boosted CBHII and endoglucanase I (EGI) activity. Our results indicated that two types of AA9 LPMOs with different modularities and regioselectivities varied in cellulose adsorption, H2O2-driven activity and synergy with cellulase on celluloses of different crystallinity which could complement each other in lignocellulose degradation. C4-oxidizing AA9 LPMOs with CBM1 were particularly essential in cellulase cocktail due to high H2O2-driven activity and a striking boosting effect on CBHI.