Structural Superimposition

Abstact

Trans-aconitate decarboxylase (Tad1) catalyzes the conversion of trans-aconitate to itaconate, a compound of growing interest in biotechnology owing to its diverse applications as a platform chemical. Despite this potential, detailed biochemical and mechanistic insights into Tad1 remain incomplete. This study presents the heterologous expression, purification, and comprehensive biochemical characterization of Tad1 from Ustilago maydis. We determined its optimal pH and temperature, and cofactor requirements. Furthermore, we employed X-ray crystallography, molecular docking, and site-directed mutagenesis to elucidate key residues involved in substrate binding and catalysis, thereby establishing a molecular basis for its decarboxylase activity. Elucidating Tad1's catalytic mechanism and active site architecture is essential for overcoming current limitations in itaconate biosynthesis and advancing sustainable chemical production.