Understanding and Mitigating the Inhibitory Effects of Lytic Polysaccharide Monooxygenase on Glycoside Hydrolases at the Early Stage of Coreaction

Lytic polysaccharide monooxygenase (LPMO) assists the degradation of recalcitrant polysaccharides in the cell wall. In this study, eight LPMOs with C1-, C4-, or C1/C4-acting modes were employed to understand their interactions with various glycoside hydrolases (GHs). Dynamic viscosity and reducing sugar measurements indicated that C1-LPMO specifically enhanced cellobiohydrolase (CBH) II activity while suppressing CBH I, whereas C4-LPMO selectively promoted CBH I but inhibited CBH II, demonstrating LPMO's regioselective modulation of GHs. Fluorescence microscopy and inhibition kinetics revealed that the inhibitory effects were mainly attributable to substrate competition at the early stage of coreaction and feedback inhibition by oxidized oligosaccharides produced by LPMO. Notably, coimmobilization of BsLPMO10A from <i>Bacillus subtilis</i> with Cellic CTec3 was effective against natural biomass substrates (<i>p</i> < 0.05), compared with the mixture of free enzymes. These findings provided novel insights into understanding and mitigating the inhibitory effects of LPMO on GH, enabling efficient saccharification of lignocellulosic plant biomass.