The role of oxidative enzymes during enzymatic hydrolysis of lignocellulosic material at industrial conditions

Lytic polysaccharide monooxygenases (LPMOs, previously GH61 and CBM33) have recently received much attention due to the their synergistic actions with classical cellulases. The catalytic cycle of LPMOs requires molecular oxygen and two electrons to complete the cycle. These electrons have in many model studies been provided by either a chemical reductant, such as ascorbate, or an enzyme, such as cellobiose dehydrogenase. Most current commercial cellulose preparations include LPMOs but questions still remain open regarding the action of these enzymes during hydrolysis of lignocellulosic materials and how it is influenced by industrial scale operating conditions.  Results from our studies using commercial cellulose preparations and isolated LPMOs strongly indicate that during action on lignocellulosic substrates lignin and/or lignin derived compounds act as the electron donor in the redox cycle of LPMOs. This demonstrates a possible new coupling between lignin modification and cellulose depolymerisation during plant cell wall deconstruction. Consequently, extra levels of complexity are involved in controlling the kinetics of LPMOs, namely the availability of lignin and the level of dissolved oxygen. These are factors influenced by choice of feedstock, pretreatment method and operating conditions during hydrolysis and fermentation. Using a bioreactor designed for efficient mixing of hydrolysates at up to 40% dry matter, the action of LPMOs was studied at industrial relevant conditions for ethanol production. The influence of feedstock and pretreatment conditions on LPMO activity was also investigated. Together these findings add an important piece to the puzzle regarding the potential of LPMOs in biorefineries.