초록
<P><B>Background</B></P><P><I>Aspergillus terreus</I> is a natural producer of itaconic acid and is currently used to produce itaconic acid on an industrial scale. The metabolic process for itaconic acid biosynthesis is very similar to the production of citric acid in <I>Aspergillus niger.</I> However, a key enzyme in <I>A. niger</I>, <I>cis</I>-aconitate decarboxylase, is missing. The introduction of the <I>A. terreus cadA</I> gene in <I>A. niger</I> exploits the high level of citric acid production (over 200 g per liter) and theoretically can lead to production levels of over 135 g per liter of itaconic acid in <I>A. niger</I>. Given the potential for higher production levels in <I>A. niger</I>, production of itaconic acid in this host was investigated.</P><P><B>Results</B></P><P>Expression of <I>Aspergillus terreus cis-</I>aconitate decarboxylase in <I>Aspergillus niger</I> resulted in the production of a low concentration (0.05 g/L) of itaconic acid. Overexpression of codon-optimized genes for <I>cis-</I>aconitate decarboxylase, a mitochondrial transporter and a plasma membrane transporter in an oxaloacetate hydrolase and glucose oxidase deficient <I>A. niger</I> strain led to highly increased yields and itaconic acid production titers. At these higher production titers, the effect of the mitochondrial and plasma membrane transporters was much more pronounced, with levels being 5–8 times higher than previously described.</P><P><B>Conclusions</B></P><P>Itaconic acid can be produced in <I>A. niger</I> by the introduction of the <I>A. terreus cis-</I>aconitate decarboxylase encoding <I>cadA</I> gene. This results in a low itaconic acid production level, which can be increased by codon-optimization of the <I>cadA</I> gene for <I>A. niger.</I> A second crucial requirement for efficient production of itaconic acid is the expression of the <I>A. terreus mttA</I> gene, encoding a putative mitochondrial transporter. Expression of this transporter results in a twenty-fold increase in the secretion of itaconic acid. Expression of the <I>A. terreus</I> itaconic acid cluster consisting of the <I>cadA</I> gene, the <I>mttA</I> gene and the <I>mfsA</I> gene results in <I>A. niger</I> strains that produce over twenty five-fold higher levels of itaconic acid and show a twenty-fold increase in yield compared to a strain expressing only CadA.</P>