Biochemical and structural characterization of enzymes in the 4-hydroxybenzoate catabolic pathway of lignin-degrading white-rot fungi

White-rot fungi (WRF) are the most efficient lignin degrading organisms in nature. However, their capacity to use lignin-related aromatic compounds, such as 4-hydroxybenzoate, as carbon sources has been only recently described and proposed to occur via the hydroxyquinol pathway. To date, the hydroxyquinol pathway has not been fully characterized in any single species from lignin-related aromatic compounds and information on the structure-function relationships of these enzymes is unavailable. Here, we characterize seven enzymes from two WRF, Trametes versicolor and Gelatoporia subvermispora, which constitute a four-enzyme biochemical cascade from 4-hydroxybenzoate to β-ketoadipate, via the hydroxyquinol pathway. Furthermore, we solve the crystal structure of four enzymes from this pathway and identify mechanistic differences with the closest bacterial and fungal structural homologs. Overall, this research expands our understanding of aromatic catabolism by WRF and establishes an alternative strategy for the conversion of lignin-related compounds to the valuable molecule β-ketoadipate, contributing to the development of biological processes for lignin valorization.