TY  - JOUR
AU  - Marienhagen, J.
AU  - Eggeling, L.
TI  - Metabolic Function of Corynebacterium glutamicum Aminotransferases AlaT and AvtA and Impact on L-Valine Production
JO  - Applied and environmental microbiology
VL  - 74
SN  - 0099-2240
CY  - Washington, DC [u.a.]
PB  - Soc.
M1  - PreJuSER-950
SP  - 7457 - 7462
PY  - 2008
N1  - We thank H. Sahm and M. Bott for continuous support, K. Krumbach for strain construction, and T. Bartek and C. Rudolf (IBT 2, Forschungszentrum Julich, Germany) for assistance with the SIXFORS fermentations.This work was supported by Deutsche Bundestiftung Umwelt Projekt 13158.
AB  - Aminotransferases (ATs) interacting with L-alanine are the least studied bacterial ATs. Whereas AlaT converts pyruvate to L-alanine in a glutamate-dependent reaction, AvtA is able to convert pyruvate to L-alanine in an L-valine-dependent manner. We show here that the wild type of Corynebacterium glutamicum with a deletion of either of the corresponding genes does not exhibit an explicit growth deficiency. However, a double mutant was auxotrophic for L-alanine, showing that both ATs can provide L-alanine and that they are the only ATs involved. Kinetic studies with isolated enzymes demonstrate that the catalytic efficiency, k(cat)/K(m), of AlaT is higher than 1 order of magnitude in the direction of L-alanine formation (3.5 x 10(4) M(-1) s(-1)), but no preference was apparent for AvtA, suggesting that AlaT is the principal L-alanine-supplying enzyme. This is in line with the cytosolic L-alanine concentration, which is reduced in the exponential growth phase from 95 mM to 18 mM by a deletion of alaT, whereas avtA deletion decreases the L-alanine concentration only to 76 mM. The combined data show that the presence of both ATs has subtle but obvious consequences on balancing intracellular amino acid pools in the wild type. The consequences are more obvious in an L-valine production strain where a high intracellular drain-off of the L-alanine precursor pyruvate prevails. We therefore used deletion of alaT to successfully reduce the contaminating L-alanine in extracellular accumulated L-valine by 80%.
KW  - Alanine: metabolism
KW  - Corynebacterium glutamicum: enzymology
KW  - Corynebacterium glutamicum: genetics
KW  - Corynebacterium glutamicum: growth & development
KW  - Cytosol: chemistry
KW  - Gene Deletion
KW  - Kinetics
KW  - Pyruvic Acid: metabolism
KW  - Substrate Specificity
KW  - Transaminases: genetics
KW  - Transaminases: isolation & purification
KW  - Transaminases: metabolism
KW  - Valine: biosynthesis
KW  - Pyruvic Acid (NLM Chemicals)
KW  - Alanine (NLM Chemicals)
KW  - Valine (NLM Chemicals)
KW  - Transaminases (NLM Chemicals)
KW  - J (WoSType)
LB  - PUB:(DE-HGF)16
C6  - pmid:18931286
C2  - pmc:PMC2607172
UR  - <Go to ISI:>//WOS:000261513700001
DO  - DOI:10.1128/AEM.01025-08
UR  - https://juser.fz-juelich.de/record/950
ER  -