Organic acid pathway
The invention relates to the use of a cytosolic citric acid synthase for the heterologous production of citrate outside the mitochondrion of a micro-organism or algae, wherein the protein is selected from A. niger An08g10920, An01g09940, An09g03570, or an ortholog of these genes. Such production is achieved by introducing the nucleic acid encoding such a protein into a suitable host cell. Preferably the protein is A. niger An08g10920, An01g09940, An09g03570 or an ortholog thereof, more particularly, wherein such an ortholog is chosen from the group of proteins listed in FIG. 9 and proteins having a percentage identity of 70%, more preferably 75%, more preferably 80%, more preferably 85%, more preferably 90%, more preferably 95%, more preferably 98%, more preferably 99% with An08g10920, An01g09940 or An09g03570.
1. A method for producing itaconic acid which method comprises modifying a eukaryotic host cell wherein said modifying consists of providing said host cell with both a nucleic acid that expresses a gene encoding a citric acid synthase that lacks a signal for expression in the mitochondrion, wherein the citric acid synthase is that encoded by an A. niger gene designated An08g10920 (SEQ ID NO: 18), or An01g09940 (SEQ ID NO: 50), or An09g03570 (SEQ ID NO: 49), or is an ortholog thereof,
wherein the ortholog is a protein that has citric acid synthase activity and a percentage identity of at least 95% with that encoded by An08g10920 (SEQ ID NO: 18), or An01g09940 (SEQ ID NO: 50) or An09g03570 (SEQ ID NO: 49); and
with a nucleic acid that expresses a gene encoding cis-aconitic acid decarboxylase (CAD); and
culturing said eukaryotic host cell.
2. The method of claim 1 , wherein said modifying is effected by transforming the host cell with a vector comprising said nucleic acid that expresses sequence encoding said citric acid synthase and a vector comprising said nucleic acid that expresses sequence encoding CAD.
3. The method of claim 1 , wherein said host cell is a fungus or a yeast or a plant or algal cell.
4. The method of claim 1 , wherein said host cell is selected from the group of Aspergillus spp., Neurospora spp., Sclerotina, Gibberella, Coniothyrium, Psiticum, Magnaporthe, Podospora, Chaetomium, Phaeosphaeria, Botryotinia, Neosartorya, Pyrenophora, Panicum, Aureococcus, Penicillium, Trichoderma, Sordaria, Colleotrichum, Verticillium, Arthrobotrys, Nectria, Leptosphaeria, Fusarium, Glomerella, Geomyces, Myceliophthora, Pichia, Saccharomyces spp., Zygosaccharomyces, Schizosaccharomyces pombe, Kluyveromyces spp., Yarrowia lipolytica, Monascus spp., Penicillium spp., Hansenula spp., Torulaspora delbrueckii, Hypomyces spp., Dotatomyces spp., Issatchenko orientalis, Phoma spp., Eupenicillium spp., Gymnoascus spp., Pichia labacensis, Pichia anomala, Wickerhamomyces anomalus, Candida cariosilognicola, Paecilomyces virioti, Scopulariopsis brevicaulis, Brettanomyces spp., Dekkera bruxellensis, Dekkera anoma and Trichoderma spp.