Characterization of the Gamma-Aminobutyric Acid (GABA) Biosynthetic Gene Cluster in High GABA-Producing Enterococcus avium G-15

Abstract

We have previously shown that the lactic acid bacterium (LAB) Enterococcus avium G-15 produces gamma-aminobutyric acid (GABA) from monosodium l-glutamate (Glu) at a hyper conversion rate. We have also found a gene cluster, designated as a gad cluster, that consists of four genes for the conversion of Glu to GABA, a Glu–GABA antiporter, and two transcriptional regulatory proteins, GadR1 and GadR2. The present study has been designed to investigate what characteristics of the GadG enzyme may contribute to the high production of GABA and how these two regulators play a role in high GABA productivity. The kinetic study showed that compared with E. coli glutamate decarboxylase (GAD) enzymes, GadG has relatively high Km (1.3–2.4 times) and kcat (1.3–1.6 times) values, indicating that although there are no remarkable differences in kinetic parameters between the three GAD enzymes, GadG may contribute to the high production of GABA in the presence of enough substrates. Further, the G-15 strain lacks the ornithine decarboxylase pathway-based acid resistance mechanism observed in some LAB strains, suggesting that the GAD-based acid resistance system is relatively important and may be vigorously employed in the G-15 strain. The molecular biological analysis of GadR1 revealed that the protein plays a role in GABA production as a transcriptional activator through an indirect pathway.