Aspergillus nigeris a well-established host organism for the production of carboxylic acids. Acids like citric, gluconic and oxalic acids can already be produced by A. niger and high titers are obtained. The formation of carboxylic acids involves the shuttling of intermediate metabolites between different intracellular compartments and utilizes different enzymatic capabilities of the respective compartment. The knowledge about the involved shuttling mechanisms and the localization of the necessary enzymes is still fragmentary. Using fluorescence microscopy, it is possible to characterize the intracellular localization of GFP tagged proteins and hence mitochondrial leader sequences can be functionally tested. In order to analyze the influence of the compartmentalization on the organic acid production, we have chosen itaconic acid as a target substance. Itaconic acid, which was selected by the US Department of Energy as one of the 12 building block chemicals for the industrial biotechnology, is currently produced by A. terreus. Heterologous expression of the A. terreus cadA gene also enables the formation of itaconic acid in A. niger although only low titers are obtained. We set out to characterize the influence of the compartmentalization on the productivity and re-engineered the enzymatic cascade by flipping the enzymatic activities of the cis-aconitic acid decarboxylase and aconitase between the mitochondrion and the cytosol. We will present new leader sequences for mitochondrial targeting in A. niger alongside with results about the positive impact of the enzymatic re-localization on the itaconic acid production.
Full conference title:
27th Fungal Genetics Conference
- Fungal Genetics Conference 27th (2013)