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:Corynebacterium glutamicum shows a great potential for the production of gamma-aminobutyric acid (GABA) from glucose fermentation via putrescine. GABA, a non-protein amino acid widespread in nature, is a component of pharmaceuticals, foods and the biodegradable plastic polyamide 4. Here, the effect of GABA in the growth of C. glutamicum was evaluated. It was estimated that the presence 1.1 M of GABA in the medium reduces the maximum growth rate of C. glutamicum to half. It was also shown that the presence of GABA in the medium negatively affects the growth of C. glutamicum in ethanol as sole carbon source. Furthermore, a new route for the production of GABA in C. glutamicum was established. GABA production from glucose fermentation via putrescine was achieved by plasmid-based overexpression of putrescine transaminase (PatA) and gamma-aminobutyraldehyde dehydrogenase (PatD) in a putrescine production strain. The resultant strain can produce 5.3 ± 0.1 g L-1 of GABA. GABA production was improved by avoiding the formation of N-acetylputrescine and by reducing the amount of nitrogen in CGXII medium. Deletion of the genes responsible for GABA catabolism and GABA re-uptake led to an increase in the GABA production of 21% achieving a titer 8.0 ± 0.3 g L-1 and an increase in the volumetric productivity of 41% reaching a productivity of 0.31 g L-1 h-1, the highest volumetric productivity achieved so far for GABA production in C. glutamicum from glucose fermentation in flasks fermentations. The results obtained hitherto are very promising and competitive compared to the traditional pathway for the production of GABA.
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: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.
Project description:5-aminovalerate (5AVA), L-lysine derived compound, represents a potential building block for the production of the bio-plastic nylon-5. Escherichia coli has been engineered for the production of 5AVA, but Corynebacterium glutamicum has never been engineered for the production of 5AVA, but, a lot of work was done in the last decades to optimize the production of the precursor L-lysine and more recently cadaverine. 5AVA added to the growth medium hardly affected growth rate of C. glutamicum, since, a half-inhibitory concentration of 1.1 M 5AVA was determined. While in E. coli, 5AVA production was engineered by using the DavBA pathway from Pseudomonas putida, here a pathway based on the route described in P. aeruginosa was established. C. glutamicum wild type was converted into a 5AVA producing strain by heterologous expression of L-lysine decarboxylase (LdcC), putrescine transaminase (PatA) and γ-aminobutyraldehyde dehydrogenase (PatD) genes from E. coli. 5AVA production was improved by using a strain previously engineered for high L-lysine production, by de-repressing phosphoenolpyruvate phosphotransferase system (PTS) and glycolysis and by avoiding formation of the by-product L-lactate. 5AVA accumulation by this strain was increased to 44.9 mM, representing a yield of 202 mmol mol-1 glucose, which is about three times higher than the highest yield achieved in E. coli for the production of 5AVA from glucose fermentation.
Project description:To identify genes which are differentially expressed in Corynebacterium glutamicum chassis C1 in comparison to the prophage free strain MB001, we performed DNA microarray analyses of C. glutamicum C1 against MB001. For this purpose RNA was isolated from cells cultivated in CGXII minimal medium with 2% glucose (w v-1) and harvested in the exponential growth phase at an OD600 of 5. Four biological replicates were performed.
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: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: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: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.