Project description:Ciprofloxacin, an inhibitor of bacterial gyrase and topoisomerase IV, was shown to inhibit growth of C. glutamicum with concomitant excretion of L-glutamate. C. glutamicum strains overproducing L-lysine, L-arginine, L-ornithine, and putrescine, respectively, produced L-glutamate instead of the desired amino acid when exposed to ciprofloxacin. Even in the absence of the putative L-glutamate exporter gene yggB, ciprofloxacin effectively triggered L-glutamate production. When C. glutamicum wild type was cultivated under nitrogen-limiting conditions, 2-oxoglutarate rather than L-glutamate was produced as consequence of exposure to ciprofloxacin. Transcriptome analysis revealed that ciprofloxacin increased RNA levels of genes involved in DNA synthesis, repair and modification. Enzyme assays showed that 2-oxoglutarate dehydrogenase activity was decreased due to ciprofloxacin addition. Here, it was shown for the first time that production of L-glutamate by C. glutamicum may be triggered by an inhibitor of DNA synthesis and L-glutamate titers of up to 37 ± 1 mM and a substrate specific L-glutamate yield of 0.13 g/g were reached.
Project description:Profiles of two major acyl-modifications, lysine acetylation and succinylation, under L-glutamate-producting and non-producing conditions in Corynebacterium glutamicum, which is industrially utilized for amino acid fermentation, was analyzed. During glutamate overproduction induced by Tween 40, global lysine acetylation was decreased, while lysine succinylation was increased. A label-free semi-quantitative proteomic analysis identified 591 acetylated proteins with 1,509 unique acetylation sites and 297 succinylated proteins with 790 unique succinylation sites. Lysine acetylation and succinylation targeted most enzymes in the central carbon metabolic pathways that are directly related to glutamate production, including the 2-oxoglutarate dehydrogenase complex (ODHC), a key enzyme for glutamate overproduction.
Project description:Metabolically engineered Corynebacterium glutamicum strains were constructed for the enhanced production of L-arginine, and their gene expression profiles were investigated
Project description:Metabolically engineered Corynebacterium glutamicum strains were constructed for the enhanced production of L-arginine, and their gene expression profiles were investigated Gene expression profiles of two C. glutamicum strains AR2 and AR6 were examined for the 3043 genes twice.
Project description:The dicarboxylic acid glutarate is gaining attention in the chemical and pharmaceutical industry as promising building-block. Synthesis of glutarate via microbial fermentation is a desirable aim which will allow the production of biopolymers avoiding fossil raw materials. Here, by rational metabolic engineering of the biofactory microorganism Corynebacterium glutamicum the fermentative production of glutarate from glucose was established. Modifications focused on increase glucose consumption and reduce by-products formation together with the heterologous overexpression of the L-lysine decarboxylase, putrescine transaminase and putrescine dehydrogenase genes from E. coli in the L-lysine producer GRLys1 allowed production the glutarate precursor 5-aminovalerate. Additional heterologous overexpression of 5-aminovalerate amino transferase and glutarate-semialdehyde dehydrogenase genes from C. glutamicum and three Pseudomonas species enabled glutarate synthesis from glucose. By coupling glutarate production with the glutamate synthesis of C. glutamicum glutarate titer improved 10%. The final strain was tested in a glucose-based fed-batch fermentation
Project description:To identify genes which are differentially expressed in Corynebacterium glutamicum in the cg2460 deletion strain, we performed DNA microarray analyses of C. glutamicum Δcg2460 compared to the WT.
Project description:To identify genes which are differentially expressed in Corynebacterium glutamicum in the cg2699 deletion strain, we performed DNA microarray analyses of C. glutamicum Δcg2699 compared to the WT.
Project description:Recently, we established Corynebacterium glutamicum as a suitable production host for various bacteriocins including garvicin Q (GarQ). Here, we establish secretion of GarQ by C. glutamicum via the Sec translocon. At neutral pH, the cationic peptide is efficiently adsorbed to the negatively charged envelope of producer bacteria limiting availability of the bacteriocin in culture supernatants. Using a reporter strain and proteomic analyses, we identified HtrA, a protease associated with secretion stress, as another potential factor limiting GarQ production.