Molecular Analysis of Glutamate Decarboxylases in Enterococcus avium

Abstract

Enterococcus avium (E. avium) is a common bacterium inhabiting the intestines of humans and other animals. Most strains of this species can produce gamma-aminobutyric acid (GABA) via the glutamate decarboxylase (GAD) system, but the presence and genetic organization of their GAD systems are poorly characterized. In this study, our bioinformatics analyses showed that the GAD system in E. avium strains was generally encoded by three gadB genes (gadB1, gadB2, and gadB3), together with an antiporter gene (gadC) and regulator gene (gadR), and these genes are organized in a cluster. This finding contrasts with that for other lactic acid bacteria. E. avium SDMCC050406, a GABA producer isolated from human feces, was employed to investigate the contribution of the three gadB genes to GABA biosynthesis. The results showed that the relative expression level of gadB3 was higher than those of gadB1 and gadB2 in the exponential growth and stationary phases, and this was accompanied by the synchronous transcription of gadC. After heterologous expression of the three gadB genes in Escherichia coli BL21 (DE3), the Km value of the purified GAD3 was 4.26 ± 0.48 mM, a value lower than those of the purified GAD1 and GAD2. Moreover, gadB3 gene inactivation caused decreased GABA production, accompanied by a reduction in resistance to acid stress. These results indicated that gadB3 plays a crucial role in GABA biosynthesis and this property endowed the strain with acid tolerance. Our findings provided insights into how E. avium strains survive the acidic environments of fermented foods and throughout transit through the stomach and gut while maintaining cell viability.

Two putative glutamate decarboxylases of Streptococcus pneumoniae as possible antigens for the production of anti-GAD65 antibodies leading to type 1 diabetes mellitus

“…Serotype 3 strains form highly mucoid colonies and its capsular polysaccharide protects the bacterium from opsonophagocytosis (Neeleman et al 1999 (gadC) is closely located to the gadB (or gadA) gene (encoding GAD) and a transcriptional regulatorcoding gene (gadR); the gadB and gadC genes frequently lie next or near each other. However, the genetic organization of the GAD system shows species and even strain speci city in LAB and other bacterial species and the gadC and gadR genes may be either absent or located elsewhere in the chromosome (Gu et al 2021). Generally, LAB species contain a GABA antiporter, but Limosilactobacillus fermentum possesses a GAD that is not accompanied by a GABA antiporter whereas Limosilactobacillus reuteri has two GABA antiporters (Cui et al 2020).…”

Two putative glutamate decarboxylases of Streptococcus pneumoniae as possible antigens for the production of anti-GAD65 antibodies leading to type 1 diabetes mellitus

“…Typically, in LAB, the GABA antiporter-coding gene ( gadC ) is closely located to the gadB (or gadA ) gene (encoding GAD) and a transcriptional regulator-coding gene ( gadR ); the gadB and gadC genes frequently lie next or near each other. However, the genetic organization of the GAD system shows species and even strain specificity in LAB and other bacterial species, and the gadC and gadR genes may be either absent or located elsewhere in the chromosome (Gu et al 2021 ). Generally, LAB species contain a GABA antiporter, but Limosilactobacillus fermentum possesses a GAD that is not accompanied by a GABA antiporter whereas Limosilactobacillus reuteri has two GABA antiporters (Cui et al 2020 ).…”

Unraveling the Role of Metals and Organic Acids in Bacterial Antimicrobial Resistance in the Food Chain

“…Often called amino acid-dependent acid resistance systems, four distinct systems may be involved in the bacterial defense against acid damage: (a) the glutamic-acid-dependent acid resistance (GDAR) system; (b) the arginine-dependent acid resistance (ADAR) system; (c) the lysine-dependent acid resistance (LDAR) system; and (d) the ornithine-dependent acid resistance (ODAR) system [ 307 ]. The GDAR system is present in several bacteria such as E. coli , Shigella flexnerii , L. monocytogenes , Lactobacillus reuteri, and Enterococcus avium [ 308 , 309 ] and provides robust protection against extreme acid stress [ 310 , 311 ]. This system is responsible for catalyzing the conversion of protonated glutamate (Glu) to Glu/γ-aminobutyrate acid (GABA) and carbon dioxide, followed by the export of GABA through the GadC antiporter in exchange for a new extracellular Glu molecule ( Figure 7 ) [ 308 ].…”