De Novo Designed β‐Hairpin Peptides Mimicking the Copper‐Binding Histidine Brace Motif of Lytic Polysaccharide Monooxygenases

Abstract Lytic polysaccharide monooxygenases (LPMOs) are Cu‐containing enzymes that play a crucial role in lignocellulosic biomass degradation for use in biofuel production. These enzymes carry out the selective oxidation of C─H bonds in the sugar units, leading to the cleavage of the glycosidic bond. Creating LPMO mimics facilitates the study of the mechanism of action, the characterisation of the reactive species responsible for the C─H bond activation and the potential scale up for industrial application. Here, we report the design, structural and functional characterisation of two novel Cu‐binding β‐hairpin peptides, called HisPins, that mimic the Histidine‐brace site of LPMOs. Activity assays were conducted with p‐nitrophenyl‐β‐d‐glucopyranoside (PNPG) and demonstrate that the Cu‐HisPins show LPMO‐like activity on the model substrate. Furthermore, the Cu‐HisPins can also perform light‐driven oxidation of phosphoric acid swollen cellulose (PASC) in the presence of melanin, similarly to some LPMO enzymes. Hence, the Cu‐HisPins represent the first structural and functional LPMO mimics based on short, folded peptide sequences and show a light‐stimulated melanin‐mediated oxidative activity, which is unreported for any LPMO mimic characterised so far. Thus, this work paves the way to further exploration of LPMO mechanism, structure‐activity relationship and substrate scope expansion.