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In the present work, the metabolic consequences of the deletion of the methionine and cysteine biosynthesis repressor protein (McbR) in Corynebacterium glutamicum, which releases almost all enzymes of methionine biosynthesis and sulfate assimilation from transcriptional regulation (D. A. Rey, A. Pühler, and J. Kalinowski, J. Biotechnol. 103:51-65, 2003), were studied. C. glutamicum ATCC 13032 DeltamcbR showed no overproduction of methionine. Metabolome analysis revealed drastic accumulation of a single metabolite, which was not present in the wild type. It was identified by isotopic labeling studies and gas chromatography/mass spectrometry as L-homolanthionine {S-[(3S)-3-amino-3-carboxypropyl]-L-homocysteine}. The accumulation of homolanthionine to an intracellular concentration of 130 mM in the DeltamcbR strain was accompanied by an elevated intracellular homocysteine level. It was shown that cystathionine-gamma-synthase (MetB) produced homolanthionine as a side reaction. MetB showed higher substrate affinity for cysteine (Km = 260 microM) than for homocysteine (Km = 540 microM). The cell is able to cleave homolanthionine at low rates via cystathionine-beta-lyase (MetC). This cleavage opens a novel threonine-independent pathway for isoleucine biosynthesis via 2-oxobutanoate formed by MetC. In fact, the deletion mutant exhibited an increased intracellular isoleucine level. Metabolic flux analysis of C. glutamicum DeltamcbR revealed that only 24% of the O-acetylhomoserine at the entry of the methionine pathway is utilized for methionine biosynthesis; the dominating fraction is either stored as homolanthionine or redirected towards the formation of isoleucine. Deletion of metB completely prevents homolanthionine accumulation, which is regarded as an important step in the development of C. glutamicum strains for biotechnological methionine production.

journal of bacteriology

Accumulation of Homolanthionine and Activation of a Novel Pathway for Isoleucine Biosynthesis in C <i>orynebacterium glutamicum</i> McbR Deletion Strains

Accumulation of Homolanthionine and Activation of a Novel Pathway for Isoleucine Biosynthesis in C orynebacterium glutamicum McbR Deletion Strains