RESUMEN
Research on enzymes for lignocellulose biomass degradation has progressively increased in recent years due to the interest in taking advantage of this natural resource. Among these enzymes are the lytic polysaccharide monooxygenases (LPMOs) that oxidatively depolymerize crystalline cellulose using a reactive oxygen species generated in a reduced monocopper active site. The copper site comprises of a highly conserved histidine-brace, providing three equatorial nitrogen ligands, whereas less conserved residues close to the copper contribute to shaping and confining the site. The catalytic copper site is exposed to the solvent and to the crystalline substrates, and as so, the influence of the copper environment on LPMO properties, including the redox potential, is of great interest. In the current work, a direct electrochemical study of an LPMO (ScLPMO10C) was conducted allowing to retrieve kinetic and thermodynamic data associated with the redox transition in the catalytic centre. Moreover, two residues that do not bind to the copper but shape the copper sites were mutated, and the properties of the mutants were compared with those of the wild-type enzyme. The direct electrochemical studies, using cyclic voltammetry, yielded redox potentials in the +200 mV range, well in line with LPMO redox potentials determined by other methods. Interestingly, while the mutations hardly affected the formal redox potential of the enzyme, they drastically affected the reactivity of the copper site and enzyme functionality.