Role of the GAD system in hop tolerance of Lactobacillus brevis

In this study, we investigated the contribution of the microbial acid stress tolerance mechanism glutamate decarboxylase (GAD) system to hop tolerance and concomitant maintenance of intracellular pH (pH(in)) in Lactobacillus brevis. In L. brevis, the GAD system comprises a transcriptional regulator (Gad-tr), a glutamate gamma-aminobutyrate antiporter (GadC) and two glutamate decarboxylases (GadB(1), GadB(2)). Hop iso-alpha-acids act as ionophores, which impair cells' proton motive force. Hop-tolerant bacteria must therefore express effective mechanisms of pH maintenance such as the GAD system. To elucidate the specific roles of the two Gad isoenzymes, we examined the influence of iso-alpha-acids on the GAD system on a metabolic and transcriptional level of two L. brevis strains. Highly hop-tolerant L. brevis TMW 1.465 proved to perform better in maintenance of pH(in) in the presence of glutamate under hop stress when compared to the rather hop-sensitive strain L. brevis TMW 1.6. The transcriptional analysis unravelled the up- or downregulation of gad-tr, gadB (1) and gadC in hop-tolerant and hop-sensitive L. brevis, respectively. Since gadB (2) expression remained fairly unaltered, we concluded that L. brevis TMW 1.6 employs both Gad isoenzymes under acid stress, whereas L. brevis TMW 1.465 manages to survive with only one isoform (GadB(2)) and can consequently master additional hop stress better by inducing gadB (1) . These findings elucidate the GAD system's role in high and low tolerance to antimicrobial hop components.