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J. Mol. Micro. Biotechnol. 3: 295-300
Pyruvate Carboxylase is a Major Bottleneck for Glutamate and Lysine Production by
Corynebacterium glutamicum
Petra G. Peters-Wendisch, Bettina Schiel, Volker F. Wendisch,
Efstratios Katsoulidis, Bettina Möckel, Hermann
Sahm, and Bernhard J. Eikmanns
Corynebacterium glutamicum possesses both phosphoenolpyruvate carboxylase (PEPCx) and
pyruvate carboxylase (PCx) as anaplerotic enzymes for growth on carbohydrates. To analyze the significance of PCx
for the amino acid production by this organism, the wild-type
pyc gene, encoding PCx, was used for the
construction of defined pyc-inactive and
pyc-overexpressing strains and the glutamate, lysine and threonine
production capabilities of these recombinant strains of
C. glutamicum were tested in comparison to the respective
host strains. No PCx activity was observed in the
pyc-inactive mutants whereas the
pyc-overexpressing strains showed eight- to elevenfold higher specific PCx activity when compared to the host strains. In a
detergent-dependent glutamate production assay, the
pyc-overexpressing strain showed more than sevenfold higher,
the PCx-deficient strain about twofold lower glutamate production than the wild-type. Overexpression of the
pyc gene and thus increasing the PCx activity in a lysine-producing strain of
C. glutamicum resulted in approximately 50% higher lysine accumulation in the culture supernatant whereas inactivation of the
pyc gene led to a decrease by 60%. In a threonine-producing strain of
C. glutamicum, the overexpression of the
pyc gene led to an only 10 to 20% increase in threonine production, however, to a more than 150% increase in the production of
the threonine precursor homoserine. These results identify the anaplerotic PCx reaction as a major bottleneck
for amino acid production by C. glutamicum and show that the enzyme is an important target for the
molecular breeding of hyperproducing strains.
Full article [pdf]
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