Journal:Acta Cryst D:S2059798323004175

Structure-function studies of a novel laccase-like multicopper oxidase from Thermothelomyces thermophila provide insights into its biological role

Christos Kosinas, Anastasia Zerva, Evangelos Topakas and Maria Dimarogona ^[1]

Molecular Tour
This study presents the crystal structure of a novel laccase-like multicopper oxidase from a thermophilic fungus (Thermothelomyces thermophila), named TtLMCO1. Multicopper oxidases are biocatalysts with various biotechnological applications, such as wastewater treatment, synthesis of novel bioactive compounds, lignin degradation as well as biosensors development. This study is focused on TtLMCO1, which is capable of oxidizing both ascorbic acid and phenolic compounds and is thus functionally categorized between ascorbate oxidases and fungal laccases. , determined using an AlphaFold 2.0 model in lack of experimentally determined structures of close homologues, revealed a three-domain laccase with two copper sites, lacking the C-terminal plug observed in other asco-laccases. The refined crystal structure of TtLMCO1 contains 576 residues arranged in three cupredoxin-like domains. includes residues 11 to 134, includes residues 135-372 and includes residues 373-602. , stabilizing the relative orientation between domains A and B. Each domain (A,B,C) is colored differently, with cyan, dark red and green respectively. The disulfide bond between residues Cys28 and Cys236 is in stick representation. Copper ions and a dioxygen molecule located at are shown as brown and red spheres respectively. Glycans are depicted as cyan sticks. . A hydrogen bond is formed the main chain carbonyl oxygen of Ile16 and N2 atom of NAG (white dotted line). . A hydrogen bond is formed between OD1 atom of (white dotted line). A CH-pi interaction is formed between the .

Analysis of solvent tunnels highlighted the amino acids that are crucial for proton transfer into the trinuclear copper site. Docking simulations with substrates that are oxidized by TtLMCO1 provide evidence that substrate specificity of these metallo-proteins is not exclusively related to their redox potential but also on the architecture of the binding site and the side chain flexibility of specific amino acids. Also, our analysis points to a potential biological role of these biocatalysts, involving the production of pigments and other bioactive compounds in their natural hosts.

References

1. ↑ Kosinas C, Zerva A, Topakas E, Dimarogona M. Structure-function studies of a novel laccase-like multicopper oxidase from Thermothelomyces thermophila provide insights into its biological role. Acta Crystallogr D Struct Biol. 2023 Jul 1;79(Pt 7):641-654. PMID:37326583 doi:10.1107/S2059798323004175