초록
<P/><P>Itaconic acid is an important platform chemical that can easily be incorporated into polymers and has the potential to replace petrochemical-based acrylic or methacrylic acid. A number of microorganisms have been developed for the biosynthesis of itaconate including <I>Aspergillus terreus</I>, <I>Escherichia coli</I>, and <I>Saccharomyces cerevisiae</I>. However, the number of strains and conditions that can be tested for increased itaconate titers are currently limited because of the lack of high-throughput screening methods. Here we identified itaconate-inducible promoters and their corresponding LysR-type transcriptional regulators from <I>Yersinia pseudotuberculosis</I> and <I>Pseudomonas aeruginosa</I>. We show that the <I>Yp</I>ItcR/P<SUB><I>ccl</I></SUB> inducible system is highly inducible by itaconic acid in the model gammaproteobacterium <I>E. coli</I> and the betaproteobacterium <I>Cupriavidus necator</I> (215- and 105-fold, respectively). The kinetics and dynamics of the <I>Yp</I>ItcR/P<SUB><I>ccl</I></SUB> inducible system are investigated, and we demonstrate, that in addition to itaconate, the genetically encoded biosensor is capable of detecting mesaconate, <I>cis</I>-, and <I>trans</I>-aconitate in a dose-dependent manner. Moreover, the fluorescence-based biosensor is applied in <I>E. coli</I> to identify the optimum expression level of <I>cadA</I>, the product of which catalyzes the conversion of <I>cis</I>-aconitate into itaconate. The fluorescence output is shown to correlate well with itaconate concentrations quantified using high-performance liquid chromatography coupled with ultraviolet spectroscopy. This work highlights the potential of the <I>Yp</I>ItcR/P<SUB><I>ccl</I></SUB> inducible system to be applied as a biosensor for high-throughput microbial strain development to facilitate improved itaconate biosynthesis.</P>