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 buildi ng block L-lysine up to rates of 2.5 nmol/min/mg (dry weight). Biochem ical analyses revealed that L-methionine represses the homoserine dehy drogenase activity and reduces the intracellular L-threonine level fro m 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 t ype of flux control at the branch point of aspartate semialdehyde conv ersion to either L-lysine or L-threonine, probably due to the absence of isoenzymes in C. glutamicum. The inducible system of L-lysine excre tion discovered was used to isolate mutants defective in the excretion of this amino acid. One such mutant characterized in detail accumulat ed 174 mM c-lysine in its cytosol without extracellular excretion of L -lysine, whereas the wild type accumulated 53 mM L-lysine in the cytos ol and 5.9 mM L-lysine in the medium. The mutant was unaffected in L-l ysine uptake or L-isoleucine or L-glutamate excretion, and also the me mbrane potential was unaltered. This mutant therefore represents a str ain with a defect in an excretion system for the primary metabolite L- lysine.