Evaluation of γ-Aminobutyric Acid (GABA) Production by Lactic Acid Bacteria Using 5-L Fermentor

Aim: This study aimed at optimizing γ-aminobutyric acid (GABA) production using lactic acid bacteria (LAB) of an Indonesian indigenous fermented buffalo milk (dadih) origin. This study utilized LAB previously cultured from dadih that has the ability to produce GABA. Materials and Methods: The study started with the identification of selected LAB by 16S rRNA, followed by optimization of GABA production by culture conditions using different initial pH, temperature, glutamate concentration, incubation time, carbon, and nitrogen sources. 16S rRNA polymerase chain reaction and analysis by phylogenetic were used to identify Lactobacillus plantarum (coded as N5) responsible for the production of GABA. Results: GABA production by high-performance liquid chromatography was highest at pH of 5.5, temperature of 36°C, glutamate concentration of 500 mM, and incubation time of 84 h. Peptone and glucose served as the nitrogen and carbon sources, respectively, whereas GABA was produced at optimum fermentation condition of 211.169 mM. Conclusion: Production of GABA by L. plantarum N5 was influenced by initial pH of 5.5, glutamic acid concentration, nitrogen source, glucose as carbon source, and incubation temperature and time. Keywords: fermented buffalo milk, Indonesian indigenous product, lactic acid bacteria, γ-aminobutyric acid.

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Production of γ-aminobutyric acid (GABA) by lactic acid bacteria strains isolated from traditional, starter-free dairy products made of raw milk

Beneficial Microbes ◽

10.3920/bm2018.0176 ◽

2019 ◽

Vol 10 (5) ◽

pp. 579-587 ◽

Cited By ~ 6

Author(s):

J.A. Valenzuela ◽

A.B. Flórez ◽

L. Vázquez ◽

O.M. Vasek ◽

B. Mayo

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Dairy Products ◽

Culture Media ◽

Monosodium Glutamate ◽

Lactobacillus Brevis ◽

Raw Milk ◽

Starter Cultures ◽

Aminobutyric Acid ◽

Gaba Production

γ-Aminobutyric acid (GABA), an amino acid not used in protein synthesis, intervenes in several physiological functions and has both diuretic and calming effects in humans. Lactic acid bacteria (LAB) strains that produce GABA could be exploited for the manufacture of health-promoting GABA-enriched dairy products. In this study, 262 LAB strains isolated from traditional dairy products made from raw milk without starter cultures were screened for GABA production in culture media supplemented with 1% monosodium glutamate (MSG) using an enzymatic (GABase) method. About half of the strains (123) were found to be GABA producers. Of these, 24, among which were 16 Lactococcus lactis subsp. lactis and three Streptococcus thermophilus strains, produced >1 mM of GABA (range 1.01-2.81 mM) and were selected for further characterisation. GABA production was confirmed in most strains by culturing in 5 mM MSG followed by HPLC quantification. A majority of the strains were confirmed to be GABA producers by this method, although lower production levels were recorded. Using species-specific primers, the gene encoding glutamate decarboxylase (GAD) was PCR-amplified in all but one of the GABA producers analysed. Amplicons sequences were compared to one another and to those held in databases. Except for one Lactobacillus brevis strain, none of the 24 GABA producers investigated produced toxic biogenic amines, such as tyramine, histamine or cadaverine. They were therefore considered safe. Either alone, in mixtures, or in combination with industrial starter or adjunct cultures, these strains might be useful in the development of health-oriented dairy products.

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Production of Gamma-Aminobutyric Acid from Lactic Acid Bacteria: A Systematic Review

International Journal of Molecular Sciences ◽

10.3390/ijms21030995 ◽

2020 ◽

Vol 21 (3) ◽

pp. 995 ◽

Cited By ~ 18

Author(s):

Yanhua Cui ◽

Kai Miao ◽

Siripitakyotin Niyaphorn ◽

Xiaojun Qu

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Genetic Engineering ◽

Molecular Mechanisms ◽

Aminobutyric Acid ◽

Research Progress ◽

Gamma Aminobutyric Acid ◽

Culture Methods ◽

Gaba Production ◽

Gaba Biosynthesis

Gamma-aminobutyric acid (GABA) is widely distributed in nature and considered a potent bioactive compound with numerous and important physiological functions, such as anti-hypertensive and antidepressant activities. There is an ever-growing demand for GABA production in recent years. Lactic acid bacteria (LAB) are one of the most important GABA producers because of their food-grade nature and potential of producing GABA-rich functional foods directly. In this paper, the GABA-producing LAB species, the biosynthesis pathway of GABA by LAB, and the research progress of glutamate decarboxylase (GAD), the key enzyme of GABA biosynthesis, were reviewed. Furthermore, GABA production enhancement strategies are reviewed, from optimization of culture conditions and genetic engineering to physiology-oriented engineering approaches and co-culture methods. The advances in both the molecular mechanisms of GABA biosynthesis and the technologies of synthetic biology and genetic engineering will promote GABA production of LAB to meet people’s demand for GABA. The aim of the review is to provide an insight of microbial engineering for improved production of GABA by LAB in the future.

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Evaluation of factors that influence the L-glutamic and γ-aminobutyric acid production duringHericium erinaceusfermentation by lactic acid bacteria

CyTA - Journal of Food ◽

10.1080/19476337.2015.1042525 ◽

2015 ◽

Vol 14 (1) ◽

pp. 47-54 ◽

Cited By ~ 18

Author(s):

Sasimar Woraharn ◽

Narissara Lailerd ◽

Bhagavathi Sundaram Sivamaruthi ◽

Wiwat Wangcharoen ◽

Sophon Sirisattha ◽

...

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Acid Production ◽

Aminobutyric Acid

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Functional Properties and Sustainability Improvement of Sourdough Bread by Lactic Acid Bacteria

Microorganisms ◽

10.3390/microorganisms8121895 ◽

2020 ◽

Vol 8 (12) ◽

pp. 1895

Author(s):

Vera Fraberger ◽

Claudia Ammer ◽

Konrad J. Domig

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Functional Properties ◽

Food Additives ◽

Dual Culture ◽

Bakery Products ◽

Gaba Production ◽

Sourdough Bread ◽

Increasing Demand

Preventing food spoilage without the addition of chemical food additives, while increasing functional properties of wheat-based bakery products, is an increasing demand by the consumers and a challenge for the food industry. Within this study, lactic acid bacteria (LAB) isolated from sourdough were screened in vitro for the ability to utilize the typical wheat carbohydrates, for their antimicrobial and functional properties. The dual culture overlay assay revealed varying levels of inhibition against the examined fungi, with Lactiplantibacillus plantarum S4.2 and Lentilactobacillusparabuchneri S2.9 exhibiting the highest suppression against the indicator strains Fusarium graminearum MUCL43764, Aspergillus fumigatus, A. flavus MUCL11945, A. brasiliensis DSM1988, and Penicillium roqueforti DSM1079. Furthermore, the antifungal activity was shown to be attributed mainly to the activity of acids produced by LAB. The antibacillus activity was evaluated by the spot-on-the-lawn method revealing a high inhibition potential of the majority of LAB isolated from sourdough against Bacillus cereus DSM31, B. licheniformis DSM13, B. subtilis LMG7135, and B. subtilis S15.20. Furthermore, evaluating the presence of the glutamate decarboxylase gen in LAB isolates by means of PCR showed a strain dependency of a potential GABA production. Finally, due to improved functional activities, LAB isolated from sourdoughs exhibit promising characteristics for the application as natural preservatives in wheat-based bakery products.

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GlnR Negatively Regulates Glutamate-Dependent Acid Resistance in Lactobacillus brevis

Applied and Environmental Microbiology ◽

10.1128/aem.02615-19 ◽

2020 ◽

Vol 86 (7) ◽

Author(s):

Luchan Gong ◽

Cong Ren ◽

Yan Xu

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Glutamate Decarboxylase ◽

Wild Type Strain ◽

Acid Resistance ◽

Lactobacillus Brevis ◽

Deletion Strain ◽

Content Type ◽

Gaba Production ◽

Encoding Gene

ABSTRACT Lactic acid bacteria often encounter a variety of multiple stresses in their natural and industrial fermentation environments. The glutamate decarboxylase (GAD) system is one of the most important acid resistance systems in lactic acid bacteria. In this study, we demonstrated that GlnR, a nitrogen regulator in Gram-positive bacteria, directly modulated γ-aminobutyric acid (GABA) conversion from glutamate and was involved in glutamate-dependent acid resistance in Lactobacillus brevis. The glnR deletion strain (ΔglnR mutant) achieved a titer of 284.7 g/liter GABA, which is 9.8-fold higher than that of the wild-type strain. The cell survival of the glnR deletion strain was significantly higher than that of the wild-type strain under the condition of acid challenge and was positively correlated with initial glutamate concentration and GABA production. Quantitative reverse transcription-PCR assays demonstrated that GlnR inhibited the transcription of the glutamate decarboxylase-encoding gene (gadB), glutamate/GABA antiporter-encoding gene (gadC), glutamine synthetase-encoding gene (glnA), and specific transcriptional regulator-encoding gene (gadR) involved in gadCB operon regulation. Moreover, GABA production and glutamate-dependent acid resistance were absolutely abolished in the gadR glnR deletion strain. Electrophoretic mobility shift and DNase I footprinting assays revealed that GlnR directly bound to the 5′-untranslated regions of the gadR gene and gadCB operon, thus inhibiting their transcription. These results revealed a novel regulatory mechanism of GlnR on glutamate-dependent acid resistance in Lactobacillus. IMPORTANCE Free-living lactic acid bacteria often encounter acid stresses because of their organic acid-producing features. Several acid resistance mechanisms, such as the glutamate decarboxylase system, F1Fo-ATPase proton pump, and alkali production, are usually employed to relieve growth inhibition caused by acids. The glutamate decarboxylase system is vital for GAD-containing lactic acid bacteria to protect cells from DNA damage, enzyme inactivation, and product yield loss in acidic habitats. In this study, we found that a MerR-type regulator, GlnR, was involved in glutamate-dependent acid resistance by directly regulating the transcription of the gadR gene and gadCB operon, resulting in an inhibition of GABA conversion from glutamate in L. brevis. This study represents a novel mechanism for GlnR's regulation of glutamate-dependent acid resistance and also provides a simple and novel strategy to engineer Lactobacillus strains to elevate their acid resistance as well as GABA conversion from glutamate.

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Biodiversity andγ-Aminobutyric Acid Production by Lactic Acid Bacteria Isolated from Traditional Alpine Raw Cow’s Milk Cheeses

BioMed Research International ◽

10.1155/2015/625740 ◽

2015 ◽

Vol 2015 ◽

pp. 1-11 ◽

Cited By ~ 36

Author(s):

Elena Franciosi ◽

Ilaria Carafa ◽

Tiziana Nardin ◽

Silvia Schiavon ◽

Elisa Poznanski ◽

...

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Random Amplified Polymorphic Dna ◽

Streptococcus Thermophilus ◽

Lactobacillus Paracasei ◽

Aminobutyric Acid ◽

Cow’S Milk ◽

Rrna Gene ◽

Cow's Milk ◽

Raw Cow’S Milk

“Nostrano-cheeses” are traditional alpine cheeses made from raw cow’s milk in Trentino-Alto Adige, Italy. This study identified lactic acid bacteria (LAB) developing during maturation of “Nostrano-cheeses” and evaluated their potential to produceγ-aminobutyric acid (GABA), an immunologically active compound and neurotransmitter. Cheese samples were collected on six cheese-making days, in three dairy factories located in different areas of Trentino and at different stages of cheese ripening (24 h, 15 days, and 1, 2, 3, 6, and 8 months). A total of 1,059 LAB isolates were screened using Random Amplified Polymorphic DNA-PCR (RAPD-PCR) and differentiated into 583 clusters. LAB strains from dominant clusters (n=97) were genetically identified to species level by partial 16S rRNA gene sequencing. LAB species most frequently isolated wereLactobacillus paracasei,Streptococcus thermophilus, andLeuconostoc mesenteroides. The 97 dominant clusters were also characterized for their ability in producing GABA by high-performance liquid chromatography (HPLC). About 71% of the dominant bacteria clusters evolving during cheeses ripening were able to produce GABA. Most GABA producers wereLactobacillus paracaseibut other GABA producing species includedLactococcus lactis,Lactobacillus plantarum,Lactobacillus rhamnosus,Pediococcus pentosaceus, andStreptococcus thermophilus. NoEnterococcus faecalisorSc. macedonicusisolates produced GABA. The isolate producing the highest amount of GABA (80.0±2.7 mg/kg) was aSc. thermophilus.

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