|Engineering of sugar metabolism of Corynebacterium glutamicum for production of amino acid L-alanine under oxygen deprivation.
Appl. Microbiol. Biotechnol. 87: 159-165. 2010.
T. Jojima, M. Fujii, E. Mori, M. Inui and H. Yukawa.
Corynebacterium glutamicum was genetically engineered to produce L-alanine from sugar under oxygen
deprivation. The genes associated with production of organic acids in C. glutamicum were inactivated and the alanine dehydrogenase gene (alaD) from Lysinibacillus sphaericus was overexpressed to direct carbon flux from organic acids to alanine.
Although the alaD-expressing strain produced alanine from glucose under oxygen deprivation,
its productivity was relatively low due to retarded glucose consumption.
Homologous overexpression of the gapA gene encoding glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in the alaD-expressing strain stimulated glucose consumption and consequently improved
alanine productivity. In contrast gapA overexpression did not affect glucose consumption under aerobic conditions,
indicating that oxygen deprivation engendered inefficient regeneration
of NAD+ resulting in impaired GAPDH activity and reduced glucose consumption
in the alanine-producing strains. Inactivation of the alanine racemase
gene allowed production of L-alanine with optical purity greater than 99.5%.
The resulting strain produced 98 g l(-1) of L-alanine after 32 h in mineral
salts medium. Our results show promise for amino acid production under