Systematic manipulation of glutathione metabolism in Escherichia coli for improved glutathione production

BACKGROUND: l-glutathione (GSH) is a non-protein thiol compound with important biological properties and is widely used in pharmaceutical, food, cosmetic and health products. The cellular GSH is determined by the activity and characteristic of GSH-synthesizing enzymes, energy and precursor supply, and degradation of formed GSH. RESULTS: In this study, genes encoding enzymes related to the precursor amino acid degradation and glycogen formation as well as GSH degradation were systematically manipulated in Escherichia coli strains over-expressing gshF from Actinobacillus succinogenes. The manipulation included disrupting the precursor degradation pathways (tnaA and sdaA), eliminating l-glutathione degradation (ggt and pepT), and manipulating the intracellular ATP level (disruption of glgB). However the constructed mutants showed lower levels of GshF expression. 2-D electrophoresis was performed to elucidate the reasons for this discrepancy, and the results indicated obvious changes in central metabolism and amino acid metabolism in the penta-mutant. Fed-batch culture of the penta-mutant ZJ12345 was performed where the GshF expression level was enhanced, and both the GSH production (19.10 mM) and the yield based on added l-cysteine (0.76 mmol/mmol) were significantly increased. CONCLUSION: By interrupting the degradation pathways of l-cysteine, serine and GSH and blocking glycogen formation, the GSH production efficiency was significantly improved.