Lactococcus lactis contains only one glutamate decarboxylase gene

Abstract

Glutamate decarboxy~ase, which is associated with a glutamate-dependent acid-resistance mechanism, was purified from Lactococcus lactis subsp. lactis by a three-step procedure. The specific activity was increased about 114-fold with a yield of 16%. The N-terminal amino acid sequence of the enzyme was determined. The gene encoding this enzyme was cloned in kcherichia coli, and its nucleotide sequence was determined. The deduced amino acid sequence suggests that the enzyme is produced as a mature form (466 amino acid residues), not as a precursor protein. The subunit molecular mass of L. lactis glutamate decarboxylase was calculated to be 53926 Da. The enzyme was maximally active a t pH 4 7 and reacted only with L-glutamate among 20 a-amino acids. The apparent Km value was calculated to be 0.51 mM. The activity was stable at acidic pH values; there was no activity in the neutral pH range. A t pH 4 1 the enzyme activity was retained at temperatures up to 70 "C in 10 min incubations. L. lactis glutamate decarboxylase behaved as a single protein when the enzyme was purified. A single band corresponding to the glutamate decarboxylase gene was detected on Southern blot analysis. These data suggest that there is one glutamate decarboxylase gene in L. lactis.

Surviving the Acid Test: Responses of Gram-Positive Bacteria to Low pH

“…Southern blot analysis of a number of strains of L. lactis subsp. lactis has shown gadB to be present in a single copy (166). A constitutively expressed gene, gadR, was found upstream.…”

Characterization of Glutamate Decarboxylase from a High γ-Aminobutyric Acid (GABA)-Producer,Lactobacillus paracasei

“…It has been reported that gadB and gadC (encoding the antiporter that exchanges glutamate for GABA) form an operon structure in L. lactis. 20) At present, it is unknown whether gadC exists upstream of gadB in Lb. paracasei.…”

“…paracasei differed from that previously reported for GAD from other LAB. 18,20) The enzyme was active within a broad pH range as compared with GADs from other LAB species.…”

“…brevis. Nomura et al 20) characterized GAD and cloned the gene responsible for GAD activity in L. lactis, and found that L. lactis contained one gene encoding GAD.…”

Identification and Probiotic Characteristics of Lactococcus Strains from Plant Materials

“…lactis bv. diacetylactis 01-7 19 . Amplified products were electrophoresed using 1.5% (w/v) agarose for 20 min at 100 V. L. lactis subsp.…”