Impact of N-Terminal Histidine Methylation on Histidine-Brace Copper(II) Peptide Models of LPMOs

In this study, we report on two synthetic tripeptides, HPH and H_MePH, designed to mimic the histidine-brace coordination motif of Lytic Polysaccharide Monooxygenases (LPMOs), with H_MePH incorporating a methylated N-terminal histidine to model a common post-translational modification in fungal LPMOs. Using a combination of potentiometric, spectroscopic, and theoretical methods, we gained structural insight into the pH-dependent mononuclear copper complexes, revealing the formation of relevant histidine-brace-ligated species above pH 6.0. The same species are formed using the two peptides, but methylation prevents the formation of dimeric species that are predominant at neutral pH in the case of the complexes of the nonmethylated HPH. Catalytic activity assays using two soluble model substrates of polysaccharides showed that both peptide complexes exhibit higher oxidative activity than copper salts when histidine-brace species are formed, with HPH species being more active than H_MePH ones in most conditions. Our findings provide a comprehensive pH-dependent analysis of histidine-brace mimics and the influence of the N-terminal modification on their structural and functional features.