Improving growth properties of Corynebacterium glutamicum by implementing an iron-responsive protocatechuate biosynthesis

Corynebacterium glutamicum experiences a transient iron limitation during growth in minimal medium, which can be compensated by the external supplementation of protocatechuic acid (PCA). Although C. glutamicum is genetically equipped to form PCA from the intermediate 3-dehydroshikimate catalysed by 3-dehydroshikimate dehydratase (encoded by qsuB), PCA synthesis is not part of the native iron-responsive regulon. To obtain a strain with improved iron availability even in the absence of the expensive supplement PCA, we re-wired the transcriptional regulation of the qsuB gene and modified PCA biosynthesis and degradation. Therefore, we ushered qsuB expression into the iron-responsive DtxR regulon by replacing the native promoter of the qsuB gene by the promoter P_ripA and introduced a second copy of the P_ripA-qsuB cassette into the genome of C. glutamicum. Reduction of the degradation was achieved by mitigating expression of the pcaG and pcaH genes through a start codon exchange. The final strain C. glutamicum IRON+ showed in the absence of PCA a significantly increased intracellular Fe²⁺ availability, exhibited improved growth properties on glucose and acetate, retained a wild type-like biomass yield but did not accumulate PCA in the supernatant. For the cultivation in minimal medium C. glutamicum IRON+ represents a useful platform strain that reveals beneficial growth properties on different carbon sources without affecting the biomass yield and overcomes the need of PCA supplementation.