Enhanced in situ H2O2 production explains synergy between an LPMO with a cellulose-binding domain and a single-domain LPMO

Lytic polysaccharide monooxygenases (LPMOs) are mono-copper enzymes that catalyze oxidative depolymerization of recalcitrant substrates such as chitin or cellulose. Recent work has shown that LPMOs catalyze fast peroxygenase reactions and that, under commonly used reaction set-ups, access to in situ generated H2O2 likely limits catalysis. Based on a hypothesis that the impact of a cellulose-binding module (CBM) on LPMO activity could relate to changes in in situ H2O2 production, we have assessed the interplay between CBM-containing ScLPMO10C and its truncated form comprising the catalytic domain only (ScLPMO10C(TR)). The results show that truncation of the linker and CBM leads to elevated H2O2 production and decreased enzyme stability. Most interestingly, combining the two enzyme forms yields strong synergistic effects, which are due to the combination of high H2O2 generation by ScLPMO10C(TR) and efficient productive use of H2O2 by the full-length enzyme. Thus, cellulose degradation becomes faster, while enzyme inactivation due to off-pathway reactions with excess H2O2 is reduced. These results underpin the complexity of ascorbic acid-driven LPMO reactions and reveal a potential mechanism for how LPMOs may interact synergistically during cellulose degradation.

Automated summary

Lytic polysaccharide monooxygenases (LPMOs) are mono-copper enzymes that catalyze oxidative depolymerization of recalcitrant substrates such as chitin or cellulose. Recent research has shown that LPMOs can catalyze fast peroxygenase reactions and that the presence of a cellulose-binding module (CBM) can affect the in situ production of H2O2. This study investigated the interplay between a CBM-containing LPMO variant and a truncated form without the CBM. The results demonstrate that truncation of the CBM leads to increased H2O2 production and decreased enzyme stability. Furthermore, the combination of the two enzyme forms results in synergistic effects, improving cellulose degradation while reducing enzyme inactivation caused by off-pathway reactions with excess H2O2.