Electrochemical Insight into the Copper Redox Chemistry and H 2 O 2 and O 2 Reducing Capability of Two AA10 Lytic Polysaccharide Monooxygenases

High Resolution Image Download MS PowerPoint Slide Lytic polysaccharide monooxygenases ([L]PMOs) are copper-containing enzymes that catalyse cleavage of the glycosidic bond, a process central to microbial biomass degradation. Here, we describe electrochemical methods used to investigate the Cu 2+/1+ redox chemistry and the polysaccharide-free catalytic activity of two AA10 LPMOs: Cj AA10B from Cellvibrio japonicus and Cf AA10 from Cellulomonas fimi . Immobilisation of these enzymes on the surface of a graphite electrode allows for direct electrochemical measurements of Cu 2+/1+ redox cycling as well as the ability of both LPMOs to reduce H 2 O 2 vs O 2 . These measurements can be advantageous when compared to biological dye assays as they provide direct kinetic measurements and allow for investigation over a wider range of environmental conditions. Values of k cat and K M - are reported for H 2 O 2 and O 2 reduction by Cj AA10B and Cf AA10 from pH 5–7, with Cf AA10 consistently outperforming Cj AA10B. Both enzymes perform faster catalysis with H 2 O 2 but when comparing the affinity-coupled specificity constant ( k cat / K M ), the LPMOs perform similarly with both H 2 O 2 and O 2, suggesting both substrates are viable. We also note an increase in redox signals as pH is decreased that correlates with EPR data suggesting a second species is formed