Phosphotransferase system-mediated glucose uptake is repressed in phosphoglucoisomerase-deficient Corynebacterium glutamicum strains.

Corynebacterium glutamicum is particularly known for its industrial application in the production of amino acids. Amino acid overproduction comes along with a high NADPH demand, which is covered mainly by the oxidative part of the pentose phosphate pathway (PPP). In previous studies, the complete redirection of the carbon flux toward the PPP by chromosomal inactivation of the pgi gene, encoding the phosphoglucoisomerase, has been applied for the improvement of C. glutamicum amino acid production strains, but this was accompanied by severe negative effects on the growth characteristics. To investigate these effects in a genetically defined background, we deleted the pgi gene in the type strain C. glutamicum ATCC 13032. The resulting strain, C. glutamicum Δpgi, lacked detectable phosphoglucoisomerase activity and grew poorly with glucose as the sole substrate. Apart from the already reported inhibition of the PPP by NADPH accumulation, we detected a drastic reduction of the phosphotransferase system (PTS)-mediated glucose uptake in C. glutamicum Δpgi. Furthermore, Northern blot analyses revealed that expression of ptsG, which encodes the glucose-specific EII permease of the PTS, was abolished in this mutant. Applying our findings, we optimized l-lysine production in the model strain C. glutamicum DM1729 by deletion of pgi and overexpression of plasmid-encoded ptsG. l-Lysine yields and productivity with C. glutamicum Δpgi(pBB1-ptsG) were significantly higher than those with C. glutamicum Δpgi(pBB1). These results show that ptsG overexpression is required to overcome the repressed activity of PTS-mediated glucose uptake in pgi-deficient C. glutamicum strains, thus enabling efficient as well as fast l-lysine production.

The transcriptional regulators RamA and RamB are involved in the regulation of glycogen synthesis in Corynebacterium glutamicum.

When grown in glucose-, fructose- or sucrose-containing medium, the amino acid producer Corynebacterium glutamicum transiently accumulates large amounts of glycogen (up to 10% of its dry weight), whereas only a marginal amount of glycogen is formed during growth with acetate. This carbon-source-dependent regulation is at least partially due to transcriptional control of glgC, encoding ADP-glucose pyrophosphorylase, the first enzyme of glycogen synthesis from glucose-1-phosphate. Here, we have analysed a possible regulatory role for the transcriptional regulators RamA and RamB on glycogen content of the cells and on control of expression of glgC and of glgA, which encodes the second enzyme of glycogen synthesis, glycogen synthase. Determination of the glycogen content of RamA- and RamB-deficient C. glutamicum indicated that RamA and RamB influence glycogen synthesis positively and negatively, respectively. In accordance with the identification of putative RamA and RamB binding sites upstream of glgC and glgA, both regulators were found to bind specifically to the glgC-glgA intergenic promoter region. Promoter activity assays in wild-type and RamA- and RamB-deficient strains of C. glutamicum revealed that (i) RamA is a positive regulator of glgC and glgA, (ii) RamB is a negative regulator of glgA and (iii) neither RamA nor RamB alone is responsible for the carbon-source-dependent regulation of glycogen synthesis in C. glutamicum.