Influence of Peptide-Rich Nitrogen Sources on GAD System Activation and GABA Production in Levilactobacillus brevis CRL 2013

Abstract

γ-Aminobutyric acid (GABA) is a bioactive metabolite valued in functional foods, but itsmicrobial production is strongly influenced by nutrient availability. Levilactobacillus brevisCRL 2013 is an efficient GABA producer; however, its biosynthesis depends on culturemedium composition. In this study, integrated physiological, proteomic, and transcriptionalanalyses were applied to assess the influence of nitrogen source composition and concentration on GABA production. No extracellular GABA was detected in a chemically definedmedium containing all amino acids and glutamate (CDMg), whereas supplementation withyeast extract or Casitone restored high-level production. The highest GABA accumulation(~250 mM) was obtained in CDMg supplemented with 1% yeast extract or 2% Casitone, anda clear peptide dose-dependent effect was observed. In contrast, other protein hydrolysatesor free amino acids alone did not stimulate GABA synthesis. Proteomic analysis revealedoverexpression of the key enzyme GadB and changes in nucleotide and fatty acid pathways.Transcriptional analysis confirmed that peptide supplementation was accompanied byincreased transcription of the gadRCB–gltX operon, in agreement with GABA accumulation. Overall, these results demonstrated that peptide composition and availability arecritical determinants of GABA biosynthesis in Lv. brevis CRL 2013and provide a basis foroptimizing peptide-based media to enhance GABA formation in food fermentations.