Date Published: June 12, 2018
Publisher: Public Library of Science
Author(s): Kejin Zhuang, Yujun Jiang, Xiaohan Feng, Li Li, Fangfang Dang, Wei Zhang, Chaoxin Man, Pratyoosh Shukla.
Gamma-aminobutyric acid (GABA) is an inhibitory neurotransmitter found in the central nervous system of mammals. A range of bacterial species can synthesize GABA, including Lactobacillus plantarum of which L-monosodium glutamate (L-MSG) is an inducer of its production. In order to synthesize GABA in high concentrations, L-MSG was utilized as the single inducing factor, a chemically defined medium (CDM) was used as the fermentation substrate, with L. plantarum CGMCC 1.2437T cultured in medium supplemented with or without L-MSG. High-throughput transcriptome sequencing was used to explore the differential genes expression of bacterial cells at 36 h of fermentation, where the GABA concentration of CDM with L-MSG reached the peak value and was 7.7 times higher than that of medium without L-MSG at the same timepoint. A total of 87 genes showed significant differential expression induced by L-MSG: of these, 69 were up-regulated genes and 18 were down-regulated. The up-regulated genes were assigned to biological processes and molecular function, while the down-regulated genes covered biological process, cellular process and molecular function. Interrogation of results using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses, indicated carbohydrate metabolism, fatty acid synthesis and amino acid metabolism were closely associated with GABA synthesis induced by L-MSG. This study provides insights into L. plantarum-mediated GABA fermentation at the molecular level and will provide a new approach for further studies related to GABA production by the other Lactic acid bacteria.
Gamma-aminobutyric acid (GABA) is a non-protein amino acid with several well-characterized physiological functions . GABA can relieve hypertension and anxiety disorders as a major inhibitory neurotransmitter in the mammalian central nervous system [2,3], and can positively affect the recovery of alcohol-related symptoms . Besides, it is also a strong secretagogue of insulin from pancreatic beta cells , which can mitigate diabetic vascular complications effectively . Due to diverse beneficial bioactivities of health-related GABA, many researchers focused on how to synthesize high levels of GABA . LAB is a developing field in food, yoghourt, beverage, and dairy food, and LAB have huge potential for GABA production in the food and pharmaceutical industry [8,9]. Lactobacillus plantarum [10,11], L. brevis [10,12], Lactococcus lactis , L. paracasei , L. delbrueckii subsp.bulgaricus , and Streptococcus salivarius are generally recognized as safe sources to produce GABA in an eco-friendly way.
This study examined induced synthesis of GABA based on L-MSG added to the culture medium as the only differential factor between experimental group and control group. In order to eliminate the influence of other irrelevant factors, a culture medium with known chemical components was used [30,31]. To further support the hypothesis that L-MSG was the only inducing factors, biomass and pH of culture broth in both experimental and control groups were not significantly different while GABA accumulation showed significant differences over time. This demonstrates that L-MSG could affect the metabolic pathways associated with GABA synthesis by the bacterium without inhibiting the growth of cells. Additionally, the differences in GABA concentration were most notable at 36–48 h of the fermentation process, when cell numbers were stable and the cellular growth was not observed. The differences relating to GABA concentration was primarily associated with different fermentation times and GABA-related metabolism. Smaller differences in GABA yield between the two groups was observed with the extended fermentation time, demonstrated by decreased levels among the experimental group and increased levels among the control group. Similar results could be seen in the study of Mazzoli et al , who speculated that although without addition of L-MSG, glutamine in the medium of control group could be converted into glutamic acid in the late fermentation, then utilized by GAD to generate small amount of GABA. And it can be hypothesized that GABA-transaminase effect led to the decomposition of GABA, which then caused the decrease of GABA in the experiment group .
The present study showed that L-MSG could induce L. plantarum CGMCC 1.2437T GABA production. The potential molecular mechanism was predicted by functional gene and pathway enrichment analysis, suggesting: (1) L-MSG induced up-regulation of gadB, which facilitated GABA synthesis; (2) L-MSG induced down-regulation of gabT, which inhibited the degradation of GABA and contributed to GABA accumulation; (3) L-MSG induced conversion of glucose to succinic acid through glycolysis and pyruvate metabolism, and the succinic acid could also inhibit the catabolic pathways of GABA; (4) L-MSG could activate the synthesis of fatty acids, which increased the membrane permeability, contributed to substrata absorption, and facilitated to cell growth and metabolism. This research presents a theoretical foundation for GABA fermentation by L. plantarum at the molecular level, provides direction for further studies, and presents and approach which may be taken for related studies.