Reconstruction of the glutamate decarboxylase system in Lactococcus lactis for biosynthesis of food-grade γ-aminobutyric acid

Lyu, Changjiang; Yao, Lili; Zhu, Qi Li; Mei, Jiaqi; Cao, Yucheng; Hu, Sheng; Wang, Zhao; Huang, Jun; Mei, Lehe; Yao, Shan‐Jing; Du, Guocheng

doi:10.1007/s00253-021-11328-5

Cited by 19 publications

(15 citation statements)

References 34 publications

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“…The yjeM gene encoding a glutamate/gamma‐aminobutyrate family transporter protein was significantly upregulated in both overexpression strains, which was consistent with the intracellular glutamate concentration during acid stress (Figures 2 and 6). The L. lactis cells could maintain dynamic balance of intracellular pH by the glutamate decarboxylase system (Lyu et al., 2021). Interestingly, the gltB gene involved in glutamate biosynthesis was repressed under acid‐stress condition, indicating that glutamate mainly enters the cell through the transport pathway.…”

Section: Discussionmentioning

confidence: 99%

“…L. lactis decarboxylate glutamate to produce gamma‐aminobutyrate (GABA), and this process consumes intracellular H + , thereby raising pH i . Meanwhile, basic GABA is exported from the cells, and its basicity is slightly stronger than that of glutamate, leading to a slightly elevated extracellular pH (Lyu et al., 2021). In addition, previous studies indicated that the arginine deiminase (ADI) system could lead to an increase of ATP generation and the depletion of acidic amino acids (aspartate and glutamate), while allowing the cells to neutralize protons by producing ammonia (Wu et al., 2012; Wu et al., 2013).…”

Section: Introductionmentioning

confidence: 99%

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Comparative transcriptome analysis reveals the contribution of amino acid transporters to acid tolerance in Lactococcus lactis

Zhu,

Zhang,

Yang

et al. 2024

Food Frontiers

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Amino acid transporters are promising targets for engineering acid‐tolerant strains of Lactococcus lactis. However, the simple overexpression of transporters alone is insufficient to achieve a highly acid‐resistant phenotype. This study investigated the effects of amino acid transporters on the acid‐stress tolerance of L. lactis. Here, we first verified the contribution of amino acid transporters to acid tolerance by overexpressing the ctrA, glnP, and glnQ genes in L. lactis. Transcriptome analysis revealed that most genes associated with specific amino acid transport, pyrimidine metabolism, and functional proteins, were upregulated in the overexpression strains. Among them, arginine biosynthesis (argG and argH), amino acid transport (yjeM and azlC), and pyrimidine metabolism were considered to be the most important regulatory mechanisms. Importantly, metabolite profiling revealed that the overexpression strains had higher intracellular levels of amino acids, particularly aspartate, glutamate, and arginine at low pH, as well as higher intracellular ATP levels, which was consistent with the corresponding gene‐expression levels. Finally, the simultaneous overexpression of glnP and glnQ led to a further improvement of acid tolerance in L. lactis. This study reveals the regulatory mechanisms of amino acid transporters, and provides a novel strategy for achieving higher acid tolerance via positive tandem expression approaches.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Comparative transcriptome analysis reveals the contribution of amino acid transporters to acid tolerance in Lactococcus lactis

Zhu,

Zhang,

Yang

et al. 2024

Food Frontiers

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“…We also observed higher amino acid sequence similarity between L. brevis Gad2 and L. plantarum GadB compared to L. brevis Gad1 (84% vs. 52% sequence similarity). The C-terminal regions of L. brevis Gad2 and L. plantarum GadB form a C-plug structure that curls toward the active site (Lyu et al, 2021). The C-terminal regions are pH dependent, and can adjust the catalytic activity of the enzyme according to pH change (Shin et al, 2014), indicating the importance of C-terminal region in enzyme activity.…”

Section: Discussionmentioning

confidence: 99%

“…brevis Gad2 and L . plantarum GadB form a C-plug structure that curls toward the active site ( Lyu et al, 2021 ). The C-terminal regions are pH dependent, and can adjust the catalytic activity of the enzyme according to pH change ( Shin et al, 2014 ), indicating the importance of C-terminal region in enzyme activity.…”

Section: Discussionmentioning

confidence: 99%

Molecular evolution and population genetics of glutamate decarboxylase acid resistance pathway in lactic acid bacteria

Sezgın

Tekin²

2023

Front. Genet.

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Glutamate decarboxylase (GAD) pathway (GDP) is a major acid resistance mechanism enabling microorganisms’ survival in low pH environments. We aimed to study the molecular evolution and population genetics of GDP in Lactic Acid Bacteria (LAB) to understand evolutionary processes shaping adaptation to acidic environments comparing species where the GDP genes are organized in an operon structure (Levilactobacillus brevis) versus lack of an operon structure (Lactiplantibacillus plantarum). Within species molecular population genetic analyses of GDP genes in L. brevis and L. plantarum sampled from diverse fermented food and other environments showed abundant synonymous and non-synonymous nucleotide diversity, mostly driven by low frequency changes, distributed throughout the coding regions for all genes in both species. GAD genes showed higher level of replacement polymorphism compared to transporter genes (gadC and YjeM) for both species, and GAD genes that are outside of an operon structure showed even higher level of replacement polymorphism. Population genetic tests suggest negative selection against replacement changes in all genes. Molecular structure and amino acid characteristics analyses showed that in none of the GDP genes replacement changes alter 3D structure or charge distribution supporting negative selection against non-conservative amino acid changes. Phylogenetic and between species divergence analyses suggested adaptive protein evolution on GDP genes comparing phylogenetically distant species, but conservative evolution comparing closely related species. GDP genes within an operon structure showed slower molecular evolution and higher conservation. All GAD and transporter genes showed high codon usage bias in examined LAB species suggesting high expression and utilization of acid resistance genes. Substantial discordances between species, GAD, and transporter gene tree topologies were observed suggesting molecular evolution of GDP genes do not follow speciation events. Distribution of operon structure on the species tree suggested multiple independent gain or loss of operon structure in LABs. In conclusion, GDP genes in LABs exhibit a dynamic molecular evolutionary history shaped by gene loss, gene transfer, negative and positive selection to maintain its active role in acid resistance mechanism, and enable organisms to thrive in acidic environments.

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“…adolescentis and E. coli Nissle 1917. The amount of GABA was 415 mM with a glutamate conversion of 92-100% for the first microorganism [198] and 17.9 g/L in the case of the second [199]. Another GABA biosynthetic is Lc.…”

Section: Recombinant Probiotics As "Cell Factories" or For The Synthe...mentioning

confidence: 99%

Recombinant Probiotic Preparations: Current State, Development and Application Prospects

Khablenko

Danylenko

Yalovenko

et al. 2023

Innov Biosyst Bioeng

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The article is devoted to the latest achievements in the field of research, development, and implementation of various types of medicinal products based on recombinant probiotics. The benefits of probiotics, their modern use in medicine along with the most frequently used genera and species of probiotic microorganisms were highlighted. The medicinal and therapeutic activities of the studied probiotics were indicated. The review suggests various methods of creating recombinant probiotic microorganisms, including standard genetic engineering methods, as well as systems biology approaches and new methods of using the CRISPR-Cas system. The range of potential therapeutic applications of drugs based on recombinant probiotics was proposed. Special attention was paid to modern research on the creation of new, more effective recombinant probiotics that can be used for various therapeutic purposes. Considering the vast diversity of therapeutic applications of recombinant probiotics and ambiguous functions, their use for the potential treatment of various common human diseases (non-infectious and infectious diseases of the gastrointestinal tract, metabolic disorders, and allergic conditions) was investigated. The prospects for creating different types of vaccines based on recombinant probiotics together with the prospects for their implementation into medicine were considered. The possibilities of using recombinant probiotics in veterinary medicine, particularly for the prevention of domestic animal diseases, were reviewed. The prospects for the implementation of recombinant probiotics as vaccines and diagnostic tools for testing certain diseases as well as modeling the work of the human digestive system were highlighted. The risks of creation, application, including the issues related to the regulatory sphere regarding the use of new recombinant microorganisms, which can potentially enter the environment and cause unforeseen circumstances, were outlined.

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Reconstruction of the glutamate decarboxylase system in Lactococcus lactis for biosynthesis of food-grade γ-aminobutyric acid

Cited by 19 publications

References 34 publications

Comparative transcriptome analysis reveals the contribution of amino acid transporters to acid tolerance in Lactococcus lactis

Comparative transcriptome analysis reveals the contribution of amino acid transporters to acid tolerance in Lactococcus lactis

Molecular evolution and population genetics of glutamate decarboxylase acid resistance pathway in lactic acid bacteria

Recombinant Probiotic Preparations: Current State, Development and Application Prospects

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