Characterization of a fungal thioesterase having claisen cyclase and deacetylase activities in melanin biosynthesis

Cell wall melanins are darkly-pigmented macromolecules that contribute to pathogenicity. In fungi, melanin biosynthesis generally involves polymerization of 1,8-dihydroxynaphthalene via a 1,3,6,8-tetrahydroxynaphthalene (THN) precursor whose backbone is assembled by iterative, nonreducing polyketide synthases. Convergent routes to THN have evolved in fungi. Parallel heptaketide and hexaketide pathways utilize conventional C-terminal thioesterase/Claisen cyclase domains and discrete side-chain deacylases. In contrast, in vitro characterization of Pks1 from Colletotrichum lagenarium establishes a true THN synthase with a bifunctional thioesterase (TE) catalyzing both cyclization and deacetylation of an enzyme-bound hexaketide. Chimeric TE domains were generated by swapping lid regions of active sites between classes of melanin-related TEs to gain insight into this unprecedented catalysis of carbon-carbon bond making and breaking by a single α/β-hydrolase fold enzyme.