title: Ustilago maydis produces itaconic acid via the unusual intermediate trans-aconitate.
abstract: Itaconic acid is an important biomass-derived chemical building block but has also recently been identified as a metabolite produced in mammals, which has antimicrobial activity. The biosynthetic pathway of itaconic acid has been elucidated in the ascomycetous fungus Aspergillus terreus and in human macrophages. In both organisms itaconic acid is generated by decarboxylation of the tricarboxylic acid (TCA) cycle intermediate cis-aconitate. Here, we show that the basidiomycetous fungus Ustilago maydis uses an alternative pathway and produces itaconic acid via trans-aconitate, the thermodynamically favoured isomer of cis-aconitate. We have identified a gene cluster that contains all genes involved in itaconic acid formation. Trans-aconitate is generated from cis-aconitate by a cytosolic aconitate-Δ-isomerase (Adi1) that belongs to the PrpF family of proteins involved in bacterial propionate degradation. Decarboxylation of trans-aconitate is catalyzed by a novel enzyme, trans-aconitate decarboxylase (Tad1). Tad1 displays significant sequence similarity with bacterial 3-carboxy-cis,cis-muconate lactonizing enzymes (CMLE). This suggests that U. maydis has evolved an alternative biosynthetic pathway for itaconate production using the toxic intermediate trans-aconitate. Overexpression of a pathway-specific transcription factor (Ria1) or a mitochondrial tricarboxylic acid transporter (Mtt1) resulted in a twofold increase in itaconate yield. Therefore, our findings offer new strategies for biotechnological production of this valuable biomass-derived chemical.
authors.affiliation: iAMB - Institute of Applied Microbiology, ABBt - Aachen Biology and Biotechnology, RWTH Aachen University, Worringerweg 1, D-52074, Aachen, Germany.
authors.forename: Elena
authors.initials: E
authors.lastname: Geiser
authors.affiliation: Department of Biology, Philipps-University Marburg, Karl-von-Frisch-Straße 8, D-35032, Marburg, Germany.
authors.forename: Sandra K
authors.initials: SK
authors.lastname: Przybilla
authors.affiliation: Department of Biology, Philipps-University Marburg, Karl-von-Frisch-Straße 8, D-35032, Marburg, Germany.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Friedrich
authors.affiliation: Department of Biology, Philipps-University Marburg, Karl-von-Frisch-Straße 8, D-35032, Marburg, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Buckel
authors.affiliation: iAMB - Institute of Applied Microbiology, ABBt - Aachen Biology and Biotechnology, RWTH Aachen University, Worringerweg 1, D-52074, Aachen, Germany.
authors.forename: Nick
authors.initials: N
authors.lastname: Wierckx
authors.affiliation: iAMB - Institute of Applied Microbiology, ABBt - Aachen Biology and Biotechnology, RWTH Aachen University, Worringerweg 1, D-52074, Aachen, Germany.
authors.forename: Lars M
authors.initials: LM
authors.lastname: Blank
authors.affiliation: Department of Biology, Philipps-University Marburg, Karl-von-Frisch-Straße 8, D-35032, Marburg, Germany.|LOEWE Center for Synthetic Microbiology (SYNMIKRO), Hans-Meerwein-Straße, D-35032, Marburg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Bölker
mesh_terms: D000156:Aconitic Acid / Q000737:chemistry / Q000378:metabolism*; D053898:Biosynthetic Pathways; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D007536:Isomerism; D013386:Succinates / Q000378:metabolism*; D014588:Ustilago / Q000235:genetics / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D013386:Succinates; D000156:Aconitic Acid; C005229:itaconic acid