Screening of GABA-producing Lactobacillus fermentum, optimized fermentation for GABA production and cloning of its glutamate decarboxylase gene

A strain of lactic acid bacteria, YS2, capable of production of aminobutyric acid (GABA), was isolated from pickled vegetable juice, and was identified as Lactobacillus fermentum by 16S rDNA sequence analysis. When YS2 was grown in MRS medium containing 1% L-monosodium glutamate (MSG), GABA yield reached 4.37 g/L. The optimal growth condition for GABA production was determined as follows: sucrose as carbon source, peptone plus beef extract as nitrogen source, initial pH at 6.0. Under the condition, GABA yield reached 5.68 g/L. The gene encoding glutamate decarboxylase, gadB, was amplified by PCR, and inserted into pEASY-E1, an expression vector for E. coli. Sequence analysis showed that the GAD from YS2 had 99% identity with that of Lactobacillus fermentum F-6. GAD was expressed with N-terminal hexahistidine tag fusion, and SDS-PAGE showed a major band of approximatela 56 kDa after purification by Ni-NTA affinity chromatography. The specific activity of purified GAD reached 1.06 U/mg.