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1.
Zool Res ; 45(6): 1175-1187, 2024 Nov 18.
Article in English | MEDLINE | ID: mdl-39318125

ABSTRACT

Isocitrate dehydrogenase 2 (IDH2) and glutamate dehydrogenase 1 (GLUD1) are key enzymes involved in the production of α-ketoglutarate (α-KG), a metabolite central to the tricarboxylic acid cycle and glutamine metabolism. In this study, we investigated the impact of IDH2 and GLUD1 on early porcine embryonic development following IDH2 and GLUD1 knockdown (KD) via double-stranded RNA (dsRNA) microinjection. Results showed that KD reduced α-KG levels, leading to delayed embryonic development, decreased blastocyst formation, increased apoptosis, reduced blastomere proliferation, and pluripotency. Additionally, IDH2 and GLUD1 KD induced abnormally high levels of trimethylation of lysine 20 of histone H4 (H4K20me3) at the 4-cell stage, likely resulting in transcriptional repression of embryonic genome activation (EGA)-related genes. Notably, KD of lysine methyltransferase 5C ( KMT5C) and supplementation with exogenous α-KG reduced H4K20me3 expression and partially rescued these defects, suggesting a critical role of IDH2 and GLUD1 in the epigenetic regulation and proper development of porcine embryos. Overall, this study highlights the significance of IDH2 and GLUD1 in maintaining normal embryonic development through their influence on α-KG production and subsequent epigenetic modifications.


Subject(s)
Embryonic Development , Epigenesis, Genetic , Glutamate Dehydrogenase , Isocitrate Dehydrogenase , Parthenogenesis , Animals , Isocitrate Dehydrogenase/genetics , Isocitrate Dehydrogenase/metabolism , Swine/embryology , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Histones/metabolism , Histones/genetics , Gene Expression Regulation, Developmental , Gene Knockdown Techniques
2.
Arch Microbiol ; 206(10): 408, 2024 Sep 19.
Article in English | MEDLINE | ID: mdl-39299989

ABSTRACT

Leishmaniasis is a complex vector-borne disease caused by intracellular protozoan parasites of the Leishmania genus. It presents a significant public health challenge in tropical and subtropical regions globally. As resistance to treatment increases, managing and controlling Leishmaniasis becomes more challenging, necessitating innovative approaches. To address this challenge, our study utilized subtractive genomics and structure-based approaches to identify common drug targets and combat antimicrobial resistance (AMR) across five Leishmania species strains. The subtractive genomics approach unraveled Glutamate Dehydrogenase (GDH) as a promising drug target for treating Leishmania infections. The investigation considered established methodologies observed in analogous studies, orthologous group, and druggability tests. Multiple sequence alignment revealed conserved sequences in GDH, while phylogenetic tree analysis provided insights into the evolutionary origin and close relationships of GDH across Leishmania species. Conserved sequences in GDH along with its function in pathogenicity provided insights into the close relationships of GDH across Leishmania species. Using a structure-based approach, our study showed the molecular interactions between GDH and three ligands-Bithionol, GW5074, and Hexachlorophene-through molecular docking and 100 ns molecular dynamics (MD) simulations. GW5074 exhibited a significant affinity for GDH, as indicated by stable RMSD values, a more compact conformation, and a higher number of hydrogen bonds than Bithionol. MMPBSA analysis confirmed the superior binding energy of the GW5074-GDH complex, emphasizing its potential as a potent ligand for drug development. This comprehensive analysis identified GW5074 as a promising candidate for inhibiting GDH activities in Leishmania species, contributing to the development of effective therapeutics against Leishmania infections.


Subject(s)
Antiprotozoal Agents , Genomics , Leishmania , Molecular Docking Simulation , Phylogeny , Leishmania/drug effects , Leishmania/genetics , Leishmania/enzymology , Antiprotozoal Agents/pharmacology , Molecular Dynamics Simulation , Glutamate Dehydrogenase/genetics , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/chemistry , Glutamate Dehydrogenase/antagonists & inhibitors , Leishmaniasis/drug therapy , Leishmaniasis/parasitology , Protozoan Proteins/genetics , Protozoan Proteins/metabolism , Protozoan Proteins/chemistry , Humans , Ligands , Sequence Alignment
3.
BMC Neurol ; 24(1): 344, 2024 Sep 13.
Article in English | MEDLINE | ID: mdl-39272024

ABSTRACT

BACKGROUND: Glioma is the most common brain tumor. IDH mutations occur frequently in glioma, indicating a more favorable prognosis. We aimed to explore energy metabolism-related genes in glioma to promote the research and treatment. METHODS: Datasets were obtained from TCGA and GEO databases. Candidate genes were screened by differential gene expression analysis, then functional enrichment analysis was conducted on the candidate genes. PPI was also carried out to help determine the target gene. GSEA and DO analysis were conducted in the different expression level groups of the target gene. Survival analysis and immune cell infiltrating analysis were performed as well. RESULTS: We screened 34 candidate genes and selected GLUD1 as the target gene. All candidate genes were significantly enriched in 10 KEGG pathways and 330 GO terms. GLUD1 expression was higher in IDH-mutant samples than IDH-wildtype samples, and higher in normal samples than tumor samples. Low GLUD1 expression was related to poor prognosis according to survival analysis. Most types of immune cells were negatively related to GLUD1 expression, but monocytes and activated mast cells exhibited significantly positive correlation with GLUD1 expression. GLUD1 expression was significantly related to 119 drugs and 6 immune checkpoint genes. GLUD1 was able to serve as an independent prognostic indicator of IDH-mutant glioma. CONCLUSION: In this study, we identified an energy metabolism-related gene GLUD1 potentially contributing to favorable clinical outcomes of IDH-mutant glioma. In glioma, GLUD1 related clinical outcomes and immune landscape were clearer, and more valuable information was provided for immunotherapy.


Subject(s)
Brain Neoplasms , Energy Metabolism , Glioma , Isocitrate Dehydrogenase , Mutation , Glioma/genetics , Glioma/metabolism , Humans , Isocitrate Dehydrogenase/genetics , Brain Neoplasms/genetics , Brain Neoplasms/metabolism , Energy Metabolism/genetics , Energy Metabolism/physiology , Prognosis , Glutamate Dehydrogenase/genetics , Glutamate Dehydrogenase/metabolism
4.
J Agric Food Chem ; 72(36): 19931-19939, 2024 Sep 11.
Article in English | MEDLINE | ID: mdl-39222309

ABSTRACT

Glutamate dehydrogenases (GDH) serve as the key regulated enzyme that links protein and carbohydrate metabolism. Combined with motif reassembly and mutation, novel GDHs were designed. Motif reassembly of thermophilic GDH and malate dehydrogenase aims to overcome stability and activity tradeoff in nonaqueous systems. Structural compatibility and dynamic cooperation of the designed AaDHs were studied by molecular dynamics simulation. Furthermore, multipoint mutations improved its catalytic activity for unnatural substrates. Amino acid interaction network analysis indicated that the high density of hydrogen-bonded salt bridges is beneficial to the stability. Finally, the experimental verification determines the kinetics of AaDHs in a nonaqueous system. The activity of Aa05 was increased by 1.78-fold with ionic liquid [EMIM]BF4. This study presents the strategy of a combination of rigid motif assembly and mutations of active sites for robust dehydrogenases with high activity in the nonaqueous system, which overcomes the activity-stability tradeoff effect.


Subject(s)
Glutamate Dehydrogenase , Molecular Dynamics Simulation , Glutamate Dehydrogenase/chemistry , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Kinetics , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Protein Engineering , Enzyme Stability , Catalytic Domain , Amino Acid Motifs , Mutation
5.
Acta Trop ; 258: 107341, 2024 Oct.
Article in English | MEDLINE | ID: mdl-39097254

ABSTRACT

Giardia duodenalis is an intestinal pathogen that is found globally. Children are more susceptible and often suffer severe consequences after infection. Despite this, the health effects of this pathogen continue to be poorly understood and neglected. In Wenzhou, Zhejiang province, China, stool samples were obtained from 1032 children who were admitted to Yuying Children's Hospital. Out of these, 684 presented with diarrhea, while 348 were asymptomatic. The stool samples were screened for G. duodenali by targeting the small subunit of the ribosomal RNA (SSU rRNA) gene. Subtypes of G. duodenalis were identified via amplification of the glutamate dehydrogenase (gdh), beta-giardin (bg), and triosephosphate isomerase (tpi) genes in samples positive for the G. duodenalis. The findings indicated the presence of G. duodenalis in 0.9 % (9/1032) of the samples, with 9/684 (1.3 %) of the samples originating from children with diarrhea and none from the asymptomatic samples. All 9 samples that tested positive for G. duodenalis were determined to be of assemblage A. Of these, 6 samples were effectively genotyped at all 3 loci, resulting in the identification of 3 distinct MLGs: MLG-AII1 (n = 1), MLG-AII2 (n = 4), and MLG-AII2 (n = 1), all belonging to G. duodenalis assemblage AII. This was the first study that confirmed G. duodenalis infections in children residing in southern Zhejiang, China, with comparatively low rates of infection. The detection of G. duodenalis assemblage AII indicates a possibility of transfer from one human to another. The parasite's effect on the health of young children requires special attention and consideration.


Subject(s)
Diarrhea , Feces , Genotype , Giardia lamblia , Giardiasis , Multilocus Sequence Typing , Humans , Giardiasis/parasitology , Giardiasis/epidemiology , Giardia lamblia/genetics , Giardia lamblia/classification , Giardia lamblia/isolation & purification , China/epidemiology , Child, Preschool , Diarrhea/parasitology , Feces/parasitology , Female , Male , Infant , Child , Protozoan Proteins/genetics , Triose-Phosphate Isomerase/genetics , Phylogeny , Glutamate Dehydrogenase/genetics , DNA, Protozoan/genetics , Prevalence
6.
Sheng Wu Gong Cheng Xue Bao ; 40(8): 2513-2527, 2024 Aug 25.
Article in Chinese | MEDLINE | ID: mdl-39174468

ABSTRACT

L-lysine is an essential amino acid with broad applications in the animal feed, human food, and pharmaceutical industries. The fermentation production of L-lysine by Escherichia coli has limitations such as poor substrate utilization efficiency and low saccharide conversion rates. We deleted the global regulatory factor gene mlc and introduced heterologous genes, including the maltose phosphotransferase genes (malAP) from Bacillus subtilis, to enhance the use efficiency of disaccharides and trisaccharides. The engineered strain E. coli XC3 demonstrated improved L-lysine production, yield, and productivity, which reached 160.00 g/L, 63.78%, and 4.44 g/(L‧h), respectively. Furthermore, we overexpressed the glutamate dehydrogenase gene (gdhA) and assimilated nitrate reductase genes (BsnasBC) from B. subtilis, along with nitrite reductase genes (EcnirBD) from E. coli, in strain E. coli XC3. This allowed the construction of E. coli XC4 with a nitrate assimilation pathway. The L-lysine production, yield, and productivity of E. coli XC4 were elevated to 188.00 g/L, 69.44%, and 5.22 g/(L‧h), respectively. After optimization of the residual sugar concentration and carbon to nitrogen ratio, the L-lysine production, yield, and productivity were increased to 204.00 g/L, 72.32%, and 5.67 g/(L‧h), respectively, in a 5 L fermenter. These values represented the increases of 40.69%, 20.03%, and 40.69%, respectively, compared with those of the starting strain XC1. By engineering the substrate utilization pathway, we successfully constructed a high-yield L-lysine producing strain, laying a solid foundation for the industrial production of L-lysine.


Subject(s)
Bacillus subtilis , Escherichia coli , Fermentation , Lysine , Metabolic Engineering , Escherichia coli/genetics , Escherichia coli/metabolism , Lysine/biosynthesis , Lysine/metabolism , Bacillus subtilis/genetics , Bacillus subtilis/metabolism , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Nitrate Reductase/genetics , Nitrate Reductase/metabolism
7.
Parasite ; 31: 50, 2024.
Article in English | MEDLINE | ID: mdl-39212527

ABSTRACT

Captive and free-living wildlife serve as significant hosts for Giardia duodenalis. Asiatic black bears, valued for their economic and medicinal importance, are extensively farmed in China and also prevalent in zoos. However, studies on G. duodenalis in these animals in China are limited. Here, 218 feces samples of Asiatic black bears were collected: 36 from a zoo in Heilongjiang Province, and 182 from a farm in Fujian Province. Nested PCR of the SSU rRNA gene, followed by sequencing, was employed to determine the frequency and assemblage distribution of G. duodenalis. Positive samples underwent further analysis through multilocus genotyping (MLG) by amplifying the genes for glutamate dehydrogenase (gdh), ß-giardin (bg), and triosephosphate isomerase (tpi). Of the 218 samples, G. duodenalis was detected in 22 cases at the SSU rRNA gene locus, including three from Heilongjiang and 19 from Fujian. Three assemblages were identified: A (n = 1), B (n = 16), and E (n = 2) in Fujian; and B (n = 3) in Heilongjiang. Out of the 22 positive samples, 20, 19, and 9 were effectively amplified and sequenced across the tpi, gdh, and bg loci, respectively. Seven samples were genotyped successfully at all three loci, identifying MLG-B1 (n = 1), MLG-B2 (n = 1), and MLG-B3 (n = 1), MLG-B4 (n = 1), MLG-B5 (n = 2), and MLG-B6 (n = 1) as the six assemblage B MLGs. This study marks the first documentation of G. duodenalis in Asiatic black bears in captivity in Fujian and Heilongjiang. The identification of zoonotic assemblages A and B, along with E, underscores potential public health concerns.


Title: Prévalence et assemblages de Giardia duodenalis chez les ours noirs d'Asie (Ursus thibetanus) d'élevage et de zoos dans les provinces chinoises du Heilongjiang et du Fujian. Abstract: Les faunes captive et libre incluent des hôtes importants pour Giardia duodenalis. Les ours noirs d'Asie, appréciés pour leur importance économique et médicinale, sont couramment élevés en Chine et répandus dans les zoos. Cependant, les études sur G. duodenalis chez ces animaux en Chine sont limitées. Ici, 218 échantillons d'excréments d'ours noirs d'Asie ont été collectés, 36 dans un zoo de la province du Heilongjiang et 182 dans une ferme de la province du Fujian. La PCR imbriquée de l'ARNr SSU, suivie d'un séquençage, a été utilisée pour déterminer la fréquence et la distribution des assemblages de G. duodenalis. Les échantillons positifs ont subi une analyse plus approfondie par génotypage multilocus (MLG) en amplifiant les gènes de la glutamate déshydrogénase (gdh), de la ß-giardine (bg) et de la triosephosphate isomérase (tpi). Sur les 218 échantillons, G. duodenalis a été détecté dans 22 cas par le locus du gène de l'ARNr SSU, dont trois du Heilongjiang et 19 du Fujian. Trois assemblages ont été identifiés : A (n = 1), B (n = 16) et E (n = 2) dans le Fujian, et B (n = 3) dans le Heilongjiang. Sur les 22 échantillons positifs, 20, 19 et 9 ont été efficacement amplifiés et séquencés respectivement pour les loci tpi, gdh et bg. Sept échantillons ont été génotypés avec succès sur les trois loci, identifiant MLG-B1 (n = 1), MLG-B2 (n = 1) et MLG-B3 (n = 1), MLG-B4 (n = 1), MLG- B5 (n = 2) et MLG-B6 (n = 1) comme les six assemblages MLG B. Cette étude marque la première investigation de G. duodenalis chez les ours noirs d'Asie en captivité au Fujian et au Heilongjiang. L'identification des assemblages zoonotiques A et B, ainsi que E, souligne des problèmes potentiels de santé publique.


Subject(s)
Animals, Zoo , Feces , Giardia lamblia , Giardiasis , Ursidae , Animals , China/epidemiology , Giardia lamblia/genetics , Giardia lamblia/isolation & purification , Giardia lamblia/classification , Giardiasis/veterinary , Giardiasis/parasitology , Giardiasis/epidemiology , Animals, Zoo/parasitology , Prevalence , Ursidae/parasitology , Feces/parasitology , Genotype , Phylogeny , Triose-Phosphate Isomerase/genetics , Farms , Glutamate Dehydrogenase/genetics , DNA, Protozoan , Protozoan Proteins/genetics , Polymerase Chain Reaction/veterinary , Multilocus Sequence Typing , Cytoskeletal Proteins/genetics
8.
Sheng Wu Gong Cheng Xue Bao ; 40(6): 1882-1894, 2024 Jun 25.
Article in Chinese | MEDLINE | ID: mdl-38914498

ABSTRACT

1,4-cyclohexanedimethylamine (1,4-BAC) is an important monomer for bio-based materials, it finds wide applications in various fields including organic synthesis, medicine, chemical industry, and materials. At present, its synthesis primarily relies on chemical method, which suffer from issues such as expensive metal catalyst, harsh reaction conditions, and safety risks. Therefore, it is necessary to explore greener alternatives for its synthesis. In this study, a two-bacterium three-enzyme cascade conversion pathway was successfully developed to convert 1,4-cyclohexanedicarboxaldehyde to 1,4-cyclohexanedimethylamine. This pathway used Escherichia coli derived aminotransferase (EcTA), Saccharomyces cerevisiae derived glutamate dehydrogenase (ScGlu-DH), and Candida boidinii derived formate dehydrogenase (CbFDH). Through structure-guided protein engineering, a beneficial mutant, EcTAF91Y, was obtained, exhibiting a 2.2-fold increase in specific activity and a 1.9-fold increase in kcat/Km compared to that of the wild type. By constructing recombinant strains and optimizing reaction conditions, it was found that under the optimal conditions, a substrate concentration of 40 g/L could produce (27.4±0.9) g/L of the product, corresponding to a molar conversion rate of 67.5%±2.1%.


Subject(s)
Escherichia coli , Saccharomyces cerevisiae , Escherichia coli/metabolism , Escherichia coli/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/enzymology , Transaminases/metabolism , Transaminases/genetics , Protein Engineering , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Formate Dehydrogenases/metabolism , Formate Dehydrogenases/genetics , Candida/enzymology , Candida/metabolism , Cyclohexylamines/metabolism
9.
J Biol Chem ; 300(7): 107473, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38879007

ABSTRACT

Provision of amino acids to the liver is instrumental for gluconeogenesis while it requires safe disposal of the amino group. The mitochondrial enzyme glutamate dehydrogenase (GDH) is central for hepatic ammonia detoxification by deaminating excessive amino acids toward ureagenesis and preventing hyperammonemia. The present study investigated the early adaptive responses to changes in dietary protein intake in control mice and liver-specific GDH KO mice (Hep-Glud1-/-). Mice were fed chow diets with a wide coverage of protein contents; i.e., suboptimal 10%, standard 20%, over optimal 30%, and high 45% protein diets; switched every 4 days. Metabolic adaptations of the mice were assessed in calorimetric chambers before tissue collection and analyses. Hep-Glud1-/- mice exhibited impaired alanine induced gluconeogenesis and constitutive hyperammonemia. The expression and activity of GDH in liver lysates were not significantly changed by the different diets. However, applying an in situ redox-sensitive assay on cryopreserved tissue sections revealed higher hepatic GDH activity in mice fed the high-protein diets. On the same section series, immunohistochemistry provided corresponding mapping of the GDH expression. Cosinor analysis from calorimetric chambers showed that the circadian rhythm of food intake and energy expenditure was altered in Hep-Glud1-/- mice. In control mice, energy expenditure shifted from carbohydrate to amino acid oxidation when diet was switched to high protein content. This shift was impaired in Hep-Glud1-/- mice and consequently the spontaneous physical activity was markedly reduced in GDH KO mice. These data highlight the central role of liver GDH in the energy balance adaptation to dietary proteins.


Subject(s)
Dietary Proteins , Energy Metabolism , Glutamate Dehydrogenase , Liver , Animals , Male , Mice , Dietary Proteins/metabolism , Gluconeogenesis , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Liver/metabolism , Mice, Knockout , Nitrogen/metabolism
10.
Int J Mol Sci ; 25(10)2024 May 13.
Article in English | MEDLINE | ID: mdl-38791334

ABSTRACT

Human evolution is characterized by rapid brain enlargement and the emergence of unique cognitive abilities. Besides its distinctive cytoarchitectural organization and extensive inter-neuronal connectivity, the human brain is also defined by high rates of synaptic, mainly glutamatergic, transmission, and energy utilization. While these adaptations' origins remain elusive, evolutionary changes occurred in synaptic glutamate metabolism in the common ancestor of humans and apes via the emergence of GLUD2, a gene encoding the human glutamate dehydrogenase 2 (hGDH2) isoenzyme. Driven by positive selection, hGDH2 became adapted to function upon intense excitatory firing, a process central to the long-term strengthening of synaptic connections. It also gained expression in brain astrocytes and cortical pyramidal neurons, including the CA1-CA3 hippocampal cells, neurons crucial to cognition. In mice transgenic for GLUD2, theta-burst-evoked long-term potentiation (LTP) is markedly enhanced in hippocampal CA3-CA1 synapses, with patch-clamp recordings from CA1 pyramidal neurons revealing increased sNMDA receptor currents. D-lactate blocked LTP enhancement, implying that glutamate metabolism via hGDH2 potentiates L-lactate-dependent glia-neuron interaction, a process essential to memory consolidation. The transgenic (Tg) mice exhibited increased dendritic spine density/synaptogenesis in the hippocampus and improved complex cognitive functions. Hence, enhancement of neuron-glia communication, via GLUD2 evolution, likely contributed to human cognitive advancement by potentiating synaptic plasticity and inter-neuronal connectivity.


Subject(s)
Cognition , Glutamate Dehydrogenase , Glutamic Acid , Neuronal Plasticity , Animals , Humans , Glutamic Acid/metabolism , Cognition/physiology , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Mice , Lactic Acid/metabolism , Long-Term Potentiation , Mice, Transgenic , Pyramidal Cells/metabolism , Hippocampus/metabolism , Evolution, Molecular , Synapses/metabolism
11.
Environ Microbiol Rep ; 16(3): e13262, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38725141

ABSTRACT

Common carp (Cyprinus carpio) were fed food with different protein concentrations following different feeding regimes, which were previously shown to affect growth, nitrogen excretion and amino acid catabolism. 16S rRNA gene amplicon sequencing was performed to investigate the gut microbiota of these fish. Lower dietary protein content increased microbial richness, while the combination of demand feeding and dietary protein content affected the composition of the gut microbiota. Hepatic glutamate dehydrogenase (GDH) activity was correlated to the composition of the gut microbiota in all dietary treatments. We found that demand-fed carp fed a diet containing 39% protein had a significantly higher abundance of Beijerinckiaceae compared to other dietary groups. Network analysis identified this family and two Rhizobiales families as hubs in the microbial association network. In demand-fed carp, the microbial association network had significantly fewer connections than in batch-fed carp. In contrast to the large effects of the feeding regime and protein content of the food on growth and nitrogen metabolism, it had only limited effects on gut microbiota composition. However, correlations between gut microbiota composition and liver GDH activity showed that host physiology and gut microbiota are connected, which warrants functional studies into the role of the gut microbiota in fish physiology.


Subject(s)
Animal Feed , Bacteria , Carps , Dietary Proteins , Gastrointestinal Microbiome , RNA, Ribosomal, 16S , Animals , Carps/microbiology , Carps/growth & development , Animal Feed/analysis , RNA, Ribosomal, 16S/genetics , Dietary Proteins/metabolism , Bacteria/classification , Bacteria/genetics , Bacteria/isolation & purification , Bacteria/metabolism , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Nitrogen/metabolism , Liver/metabolism , Phylogeny , Diet/veterinary
12.
J Mol Cell Biol ; 16(4)2024 Sep 30.
Article in English | MEDLINE | ID: mdl-38587834

ABSTRACT

Glutamate dehydrogenase 1 (GLUD1) is implicated in oncogenesis. However, little is known about the relationship between GLUD1 and hepatocellular carcinoma (HCC). In the present study, we demonstrated that the expression levels of GLUD1 significantly decreased in tumors, which was relevant to the poor prognosis of HCC. Functionally, GLUD1 silencing enhanced the growth and migration of HCC cells. Mechanistically, the upregulation of interleukin-32 through AKT activation contributes to GLUD1 silencing-facilitated hepatocarcinogenesis. The interaction between GLUD1 and AKT, as well as α-ketoglutarate regulated by GLUD1, can suppress AKT activation. In addition, LIM and SH3 protein 1 (LASP1) interacts with GLUD1 and induces GLUD1 degradation via the ubiquitin-proteasome pathway, which relies on the E3 ubiquitin ligase synoviolin (SYVN1), whose interaction with GLUD1 is enhanced by LASP1. In hepatitis B virus (HBV)-related HCC, the HBV X protein (HBX) can suppress GLUD1 with the participation of LASP1 and SYVN1. Collectively, our data suggest that GLUD1 silencing is significantly associated with HCC development, and LASP1 and SYVN1 mediate the inhibition of GLUD1 in HCC, especially in HBV-related tumors.


Subject(s)
Adaptor Proteins, Signal Transducing , Carcinoma, Hepatocellular , Cytoskeletal Proteins , Glutamate Dehydrogenase , LIM Domain Proteins , Liver Neoplasms , Viral Regulatory and Accessory Proteins , Humans , LIM Domain Proteins/metabolism , LIM Domain Proteins/genetics , Liver Neoplasms/pathology , Liver Neoplasms/metabolism , Liver Neoplasms/genetics , Liver Neoplasms/virology , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Cytoskeletal Proteins/metabolism , Cytoskeletal Proteins/genetics , Carcinoma, Hepatocellular/pathology , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/metabolism , Carcinoma, Hepatocellular/virology , Adaptor Proteins, Signal Transducing/metabolism , Adaptor Proteins, Signal Transducing/genetics , Cell Line, Tumor , Viral Regulatory and Accessory Proteins/metabolism , Ubiquitin-Protein Ligases/metabolism , Ubiquitin-Protein Ligases/genetics , Carcinogenesis/metabolism , Carcinogenesis/genetics , Carcinogenesis/pathology , Animals , Hepatitis B virus/metabolism , Hepatitis B virus/physiology , Proto-Oncogene Proteins c-akt/metabolism , Gene Expression Regulation, Neoplastic , Cell Movement/genetics , Cell Proliferation , Male , Mice , Trans-Activators
13.
Int J Mol Sci ; 25(8)2024 Apr 14.
Article in English | MEDLINE | ID: mdl-38673928

ABSTRACT

There are two paralogs of glutamate dehydrogenase (GDH) in humans encoded by the GLUD1 and GLUD2 genes as a result of a recent retroposition during the evolution of primates. The two human GDHs possess significantly different regulation by allosteric ligands, which is not fully characterized at the structural level. Recent advances in identification of the GDH ligand binding sites provide a deeper perspective on the significance of the accumulated substitutions within the two GDH paralogs. In this review, we describe the evolution of GLUD1 and GLUD2 after the duplication event in primates using the accumulated sequencing and structural data. A new gibbon GLUD2 sequence questions the indispensability of ancestral R496S and G509A mutations for GLUD2 irresponsiveness to GTP, providing an alternative with potentially similar regulatory features. The data of both GLUD1 and GLUD2 evolution not only confirm substitutions enhancing GLUD2 mitochondrial targeting, but also reveal a conserved mutation in ape GLUD1 mitochondrial targeting sequence that likely reduces its transport to mitochondria. Moreover, the information of GDH interactors, posttranslational modification and subcellular localization are provided for better understanding of the GDH mutations. Medically significant point mutations causing deregulation of GDH are considered from the structural and regulatory point of view.


Subject(s)
Evolution, Molecular , Glutamate Dehydrogenase , Protein Processing, Post-Translational , Animals , Humans , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Glutamate Dehydrogenase/chemistry , Ligands , Mutation , Primates/genetics
14.
Nat Commun ; 15(1): 3468, 2024 Apr 24.
Article in English | MEDLINE | ID: mdl-38658571

ABSTRACT

Metabolism has recently emerged as a major target of genes implicated in the evolutionary expansion of human neocortex. One such gene is the human-specific gene ARHGAP11B. During human neocortex development, ARHGAP11B increases the abundance of basal radial glia, key progenitors for neocortex expansion, by stimulating glutaminolysis (glutamine-to-glutamate-to-alpha-ketoglutarate) in mitochondria. Here we show that the ape-specific protein GLUD2 (glutamate dehydrogenase 2), which also operates in mitochondria and converts glutamate-to-αKG, enhances ARHGAP11B's ability to increase basal radial glia abundance. ARHGAP11B + GLUD2 double-transgenic bRG show increased production of aspartate, a metabolite essential for cell proliferation, from glutamate via alpha-ketoglutarate and the TCA cycle. Hence, during human evolution, a human-specific gene exploited the existence of another gene that emerged during ape evolution, to increase, via concerted changes in metabolism, progenitor abundance and neocortex size.


Subject(s)
GTPase-Activating Proteins , Glutamate Dehydrogenase , Neocortex , Neocortex/metabolism , Neocortex/embryology , Neocortex/growth & development , Neocortex/cytology , Humans , Animals , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , GTPase-Activating Proteins/metabolism , GTPase-Activating Proteins/genetics , Ketoglutaric Acids/metabolism , Neuroglia/metabolism , Glutamic Acid/metabolism , Mitochondria/metabolism , Mitochondria/genetics , Mice , Citric Acid Cycle/genetics , Female
15.
Microb Biotechnol ; 17(3): e14429, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38483038

ABSTRACT

Glutamate serves as the major cellular amino group donor. In Bacillus subtilis, glutamate is synthesized by the combined action of the glutamine synthetase and the glutamate synthase (GOGAT). The glutamate dehydrogenases are devoted to glutamate degradation in vivo. To keep the cellular glutamate concentration high, the genes and the encoded enzymes involved in glutamate biosynthesis and degradation need to be tightly regulated depending on the available carbon and nitrogen sources. Serendipitously, we found that the inactivation of the ansR and citG genes encoding the repressor of the ansAB genes and the fumarase, respectively, enables the GOGAT-deficient B. subtilis mutant to synthesize glutamate via a non-canonical fumarate-based ammonium assimilation pathway. We also show that the de-repression of the ansAB genes is sufficient to restore aspartate prototrophy of an aspB aspartate transaminase mutant. Moreover, in the presence of arginine, B. subtilis mutants lacking fumarase activity show a growth defect that can be relieved by aspB overexpression, by reducing arginine uptake and by decreasing the metabolic flux through the TCA cycle.


Subject(s)
Ammonium Compounds , Fumarate Hydratase/genetics , Glutamic Acid/metabolism , Glutamate Dehydrogenase/genetics , Arginine , Nitrogen/metabolism
16.
Mol Biol Rep ; 51(1): 403, 2024 Mar 08.
Article in English | MEDLINE | ID: mdl-38457002

ABSTRACT

BACKGROUND: Giardia duodenalis is an important intestinal parasitic protozoan that infects several vertebrates, including humans. Cattle are considered the major source of giardiasis outbreak in humans. This study aimed to investigate the prevalence and multilocus genotype (MLG) of G. duodenalis in Shanxi, and lay the foundation for the prevention and control of Giardiosis. METHODS AND RESULTS: DNA extraction, nested polymerase chain reaction, sequence analysis, MLG analysis, and statistical analysis were performed using 858 bovine fecal samples from Shanxi based on three gene loci: ß-giardin (bg), glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi). The overall prevalence of G. duodenalis was 28.3%, while its prevalence in Yingxian and Lingqiu was 28.1% and 28.5%, respectively. The overall prevalence of G. duodenalis in dairy cattle and beef cattle was 28.0% and 28.5%, respectively. G. duodenalis infection was detected in all age groups evaluated in this study. The overall prevalence of G. duodenalis in diarrhea and nondiarrhea samples was 32.4% and 27.5%, respectively, whereas that in intensively farmed and free-range cattle was 35.0% and 19.9%, respectively. We obtained 83, 53, and 59 sequences of bg, gdh, and tpi in G. duodenalis, respectively. Moreover, assemblage A (n = 2) and assemblage E (n = 81) by bg, assemblage A (n = 1) and assemblage E (n = 52) by gdh, and assemblage A (n = 2) and assemblage E (n = 57) by tpi were identified. Multilocus genotyping yielded 29 assemblage E MLGs, which formed 10 subgroups. CONCLUSIONS: To the best of our knowledge, this is the first study to report cattle infected with G. duodenalis in Shanxi, China. Livestock-specific G. duodenalis assemblage E was the dominant assemblage genotype, and zoonotic sub-assemblage AI was also detected in this region.


Subject(s)
Giardia lamblia , Giardiasis , Humans , Cattle , Animals , Giardia lamblia/genetics , Multilocus Sequence Typing , Protozoan Proteins/genetics , Giardiasis/epidemiology , Giardiasis/veterinary , Giardiasis/parasitology , Genotype , China/epidemiology , Prevalence , Feces/parasitology , Triose-Phosphate Isomerase/genetics , Glutamate Dehydrogenase/genetics , Phylogeny
17.
Cancer Res ; 84(10): 1643-1658, 2024 May 15.
Article in English | MEDLINE | ID: mdl-38417136

ABSTRACT

Hepatocellular carcinoma (HCC) is a typical tumor that undergoes metabolic reprogramming, differing from normal liver tissue in glucose, lipid, nucleic acid, and amino acid metabolism. Although ammonia is a toxic metabolic by-product, it has also been recently recognized as a signaling molecule to activate lipid metabolism, and it can be a nitrogen source for biosynthesis to support tumorigenesis. In this study, we revealed that ß-catenin activation increases ammonia production in HCC mainly by stimulating glutaminolysis. ß-Catenin/LEF1 activated the transcription of the glutamate dehydrogenase GLUD1, which then promoted ammonia utilization to enhance the production of glutamate, aspartate, and proline as evidenced by 15NH4Cl metabolic flux. ß-Catenin/TCF4 induced the transcription of SLC4A11, an ammonia transporter, to excrete excess ammonia. SLC4A11 was upregulated in HCC tumor tissues, and high SLC4A11 expression was associated with poor prognosis and advanced disease stages. Loss of SLC4A11 induced HCC cell senescence in vitro by blocking ammonia excretion and reduced ß-catenin-driven tumor growth in vivo. Furthermore, elevated levels of plasma ammonia promoted the progression of ß-catenin mutant HCC, which was impeded by SLC4A11 deficiency. Downregulation of SLC4A11 led to ammonia accumulation in tumor interstitial fluid and decreased plasma ammonia levels in HCC with activated ß-catenin. Altogether, this study indicates that ß-catenin activation reprograms ammonia metabolism and that blocking ammonia excretion by targeting SLC4A11 could be a promising approach to induce senescence in ß-catenin mutant HCC. SIGNIFICANCE: Ammonia metabolism reprogramming mediated by aberrant activation of ß-catenin induces resistance to senescence in HCC and can be targeted by inhibiting SLC4A11 as a potential therapy for ß-catenin mutant liver cancer.


Subject(s)
Ammonia , Carcinoma, Hepatocellular , Cellular Senescence , Liver Neoplasms , beta Catenin , Animals , Humans , Male , Mice , Ammonia/metabolism , beta Catenin/metabolism , Carcinoma, Hepatocellular/metabolism , Carcinoma, Hepatocellular/pathology , Carcinoma, Hepatocellular/genetics , Cell Line, Tumor , Gene Expression Regulation, Neoplastic , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Liver Neoplasms/genetics , Mice, Nude , Prognosis
18.
J Microbiol Biotechnol ; 34(4): 978-984, 2024 Apr 28.
Article in English | MEDLINE | ID: mdl-38379308

ABSTRACT

Genome-scale metabolic model (GEM) can be used to simulate cellular metabolic phenotypes under various environmental or genetic conditions. This study utilized the GEM to observe the internal metabolic fluxes of recombinant Escherichia coli producing gamma-aminobutyric acid (GABA). Recombinant E. coli was cultivated in a fermenter under three conditions: pH 7, pH 5, and additional succinic acids. External fluxes were calculated from cultivation results, and internal fluxes were calculated through flux optimization. Based on the internal flux analysis, glycolysis and pentose phosphate pathways were repressed under cultivation at pH 5, even though glutamate dehydrogenase increased GABA production. Notably, this repression was halted by adding succinic acid. Furthermore, proper sucA repression is a promising target for developing strains more capable of producing GABA.


Subject(s)
Escherichia coli , gamma-Aminobutyric Acid , Escherichia coli/genetics , Escherichia coli/metabolism , gamma-Aminobutyric Acid/metabolism , gamma-Aminobutyric Acid/biosynthesis , Hydrogen-Ion Concentration , Fermentation , Glycolysis , Succinic Acid/metabolism , Pentose Phosphate Pathway , Metabolic Flux Analysis , Models, Biological , Bioreactors/microbiology , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Metabolic Engineering/methods
19.
Microb Pathog ; 188: 106565, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38309311

ABSTRACT

Streptococcus suis serotype 2 is a zoonotic agent that causes substantial economic losses to the swine industry and threatens human public health. Factors that contribute to its ability to cause disease are not yet fully understood. Glutamate dehydrogenase (GDH) is an enzyme found in living cells and plays vital roles in cellular metabolism. It has also been shown to affect pathogenic potential of certain bacteria. In this study, we constructed a S. suis serotype 2 GDH mutant (Δgdh) by insertional inactivation mediated by a homologous recombination event and confirmed loss of expression of GDH in the mutant by immunoblot and enzyme activity staining assays. Compared with the wild type (WT) strain, Δgdh displayed a different phenotype. It exhibited impaired growth in all conditions evaluated (solid and broth media, increased temperature, varying pH, and salinity) and formed cells of reduced size. Using a swine infection model, pigs inoculated with the WT strain exhibited fever, specific signs of disease, and lesions, and the strain could be re-isolated from the brain, lung, joint fluid, and blood samples collected from the infected pigs. Pigs inoculated with the Δgdh strain did not exhibit any clinical signs of disease nor histologic lesions, and the strain could not be re-isolated from any of the tissues nor body fluid sampled. The Δgdh also showed a decreased level of survival in pig blood. Taken together, these results suggest that the gdh is important in S. suis physiology and its ability to colonize, disseminate, and cause disease.


Subject(s)
Streptococcal Infections , Streptococcus suis , Swine Diseases , Swine , Animals , Humans , Virulence , Glutamate Dehydrogenase/genetics , Glutamate Dehydrogenase/metabolism , Streptococcus suis/genetics , Serogroup , Virulence Factors/genetics , Virulence Factors/metabolism , Swine Diseases/microbiology , Streptococcal Infections/veterinary , Streptococcal Infections/microbiology
20.
J Neurochem ; 168(5): 719-727, 2024 05.
Article in English | MEDLINE | ID: mdl-38124277

ABSTRACT

The excitatory neurotransmitter glutamate has a role in neuronal migration and process elongation in the central nervous system (CNS). The effects of chronic glutamate hyperactivity on vesicular and protein transport within CNS neurons, that is, processes necessary for neurite growth, have not been examined previously. In this study, we measured the effects of lifelong hyperactivity of glutamate neurotransmission on axoplasmic transport in CNS neurons. We compared wild-type (wt) to transgenic (Tg) mice over-expressing the glutamate dehydrogenase gene Glud1 in CNS neurons and exhibiting increases in glutamate transmitter formation, release, and synaptic activation in brain throughout the lifespan. We found that Glud1 Tg as compared with wt mice exhibited increases in the rate of anterograde axoplasmic transport in neurons of the hippocampus measured in brain slices ex vivo, and in olfactory neurons measured in vivo. We also showed that the in vitro pharmacologic activation of glutamate synapses in wt mice led to moderate increases in axoplasmic transport, while exposure to selective inhibitors of ion channel forming glutamate receptors very significantly suppressed anterograde transport, suggesting a link between synaptic glutamate receptor activation and axoplasmic transport. Finally, axoplasmic transport in olfactory neurons of Tg mice in vivo was partially inhibited following 14-day intake of ethanol, a known suppressor of axoplasmic transport and of glutamate neurotransmission. The same was true for transport in hippocampal neurons in slices from Glud1 Tg mice exposed to ethanol for 2 h ex vivo. In conclusion, endogenous activity at glutamate synapses regulates and glutamate synaptic hyperactivity increases intraneuronal transport rates in CNS neurons.


Subject(s)
Glutamate Dehydrogenase , Mice, Transgenic , Neurons , Receptors, Glutamate , Animals , Mice , Glutamate Dehydrogenase/metabolism , Glutamate Dehydrogenase/genetics , Neurons/metabolism , Neurons/drug effects , Receptors, Glutamate/metabolism , Axonal Transport/drug effects , Axonal Transport/physiology , Glutamic Acid/metabolism , Hippocampus/metabolism , Mice, Inbred C57BL
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