Unbalance of L-lysine flux in Corynebacterium glutamicum and its use for the isolation of excretion-defective mutants.
We found that the simple addition of L-methionine to the wild type of Corynebacterium glutamicum results in excretion of the cellular building block L-lysine up to rates of 2.5 nmol/min/mg (dry weight). Biochemical analyses revealed that L-methionine represses the homoserine dehydrogenase activity and reduces the intracellular L-threonine level from 7 to less than 2 mM. Since L-lysine synthesis is regulated mainly by L-threonine (plus L-lysine) availability, the result is enhanced flux towards L-lysine. This indicates a delicate and not well controlled type of flux control at the branch point of aspartate semialdehyde conversion to either L-lysine or L-threonine, probably due to the absence of isoenzymes in C. glutamicum. The inducible system of L-lysine excretion discovered was used to isolate mutants defective in the excretion of this amino acid. One such mutant characterized in detail accumulated 174 mM L-lysine in its cytosol without extracellular excretion of L-lysine, whereas the wild type accumulated 53 mM L-lysine in the cytosol and 5.9 mM L-lysine in the medium. The mutant was unaffected in L-lysine uptake or L-isoleucine or L-glutamate excretion, and also the membrane potential was unaltered. This mutant therefore represents a strain with a defect in an excretion system for the primary metabolite L-lysine.
Vrljic
M
M
Biotechnologie 1, Forschungszentrum Jülich GmbH, Germany.
Kronemeyer
W
W
Sahm
H
H
Eggeling
L
L
D001692:Biological Transport / Q000235:genetics; D003352:Corynebacterium / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism; D006715:Homoserine Dehydrogenase / Q000378:metabolism; D007532:Isoleucine / Q000378:metabolism; D008239:Lysine / Q000378:metabolism*; D008715:Methionine / Q000494:pharmacology*; D009154:Mutation
D007532:Isoleucine; D018698:Glutamic Acid; D008715:Methionine; D006715:Homoserine Dehydrogenase; D008239:Lysine