Cellulose degradation by lytic polysaccharide monooxygenase fueled by an aryl-alcohol oxidase

Abstract

Enzymatic deconstruction of cellulose was dramatically changed by the discovery of Lytic Polysaccharide Monooxygenases (LPMOs). These revolutionizing enzymes are copper-dependent, and through an oxidative process, break the cellulose chain, facilitating deconstruction by cellulases. The LPMO catalytic copper can be reduced by an assortment of molecules, including lignin degradation products and other enzymes. Here we show that Thermothelomyces thermophilus LPMO9H (TtLPMO9H) had activity towards cellulosic substrate, when both an aryl-alcohol oxidase, TtAAOx, and its substrate, veratryl alcohol are present. We found that veratryl alcohol by itself could not drive the LPMO activity, but when combined with TtAAOx, the LPMO was fueled. Formation and release of oxidized oligosaccharides by TtLPMO9H required the presence of methoxylated benzyl alcohols and either TtAAOx for in situ production, or an exogenous supply of H₂O₂. Additionally, we showed that this in situ production of H₂O₂ resulted in slower LPMO reactions as compared to standard LPMO catalysis driven by ascorbic acid but circumvented early inactivation of the LPMO. These results suggest that many oxidoreductases, including oxidases associated with lignin degradation, can serve in LPMO reactions as in situ H₂O₂ generators.