Dataset Information

Molecular Characterization and Gene Expression of Glutathione Peroxidase 1 in Tor tambroides Exposed to Temperature Stress.

ABSTRACT: Temperature is an abiotic factor that affects various biological and physiological processes in fish. Temperature stress is known to increase the production of reactive oxygen species (ROS) that subsequently cause oxidative stress. Fish is known to evolve a system of antioxidant enzymes to reduce ROS toxicology. Glutathione peroxidase (GPx) family consists of key enzymes that protect fish from oxidative stress. In this study, full-length GPx1 cDNA (GenBank accession no. KY984468) of Tor tambroides was cloned and characterized by rapid amplification of cDNA ends (RACE). The 899-base-pair (bp) GPx1 cDNA includes a 576-bp open reading frame encoding for 191 amino acids, plus 28?bp of 5'-untranslated region (UTR) and 295?bp of 3'-UTR. Homology analysis revealed that GPx1 of T tambroides (Tor-GPx1) shared high similarity with GPx1 sequences of other fish species. The phylogenetic construction based on the amino acid sequence showed that Tor-GPx1 formed a clade with GPx1 sequences of various fish species. Real-time polymerase chain reaction (PCR) was performed to assess the levels of GPx1 gene expression in the liver and muscle of T tambroides under thermal stress. The results indicated that GPx1 gene expression was down-regulated under decreased temperature. However, there was no significant difference between GPx1 gene expression in fish exposed to high temperature and control. Our study provides the first data regarding GPx gene expression in T tambroides under thermal stress.

SUBMITTER: Do TD

PROVIDER: S-EPMC6572904 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Molecular Characterization and Gene Expression of Glutathione Peroxidase 1 in <i>Tor tambroides</i> Exposed to Temperature Stress.

Do Thinh Dinh TD Thi Mai Nguyen N Duy Khoa Tran Nguyen TN Abol-Munafi Ambok Bolong AB Liew Hon Jung HJ Kim Chang-Bae CB Wong Li Lian LL

Evolutionary bioinformatics online 20190613

Temperature is an abiotic factor that affects various biological and physiological processes in fish. Temperature stress is known to increase the production of reactive oxygen species (ROS) that subsequently cause oxidative stress. Fish is known to evolve a system of antioxidant enzymes to reduce ROS toxicology. Glutathione peroxidase (GPx) family consists of key enzymes that protect fish from oxidative stress. In this study, full-length GPx1 cDNA (GenBank accession no. KY984468) of <i>Tor tambr ...[more]

PMID: 31236006

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Project description:More attention has been recently directed toward glutathione peroxidase and s-transferase enzymes because of the great importance they hold with respect to their applications in the pharmaceutical field. This work was conducted to optimize the production and characterize glutathione peroxidase and glutathione s-transferase produced by Lactobacillus plantarum KU720558 using Plackett-Burman and Box-Behnken statistical designs. To assess the impact of the culture conditions on the microbial production of the enzymes, colorimetric methods were used. Following data analysis, the optimum conditions that enhanced the s-transferase yield were the De Man-Rogosa-Sharp (MRS) broth as a basal medium supplemented with 0.1% urea, 0.075% H2O2, 0.5% 1-butanol, 0.0125% amino acids, and 0.05% SDS at pH 6.0 and anaerobically incubated for 24 h at 40°C. The optimum s-transferase specific activity was 1789.5 U/mg of protein, which was ~12 times the activity of the basal medium. For peroxidase, the best medium composition was 0.17% urea, 0.025% bile salt, 7.5% Na Cl, 0.05% H2O2, 0.05% SDS, and 2% ethanol added to the MRS broth at pH 6.0 and anaerobically incubated for 24 h at 40°C. Furthermore, the optimum peroxidase specific activity was 612.5 U/mg of protein, indicating that its activity was 22 times higher than the activity recorded in the basal medium. After SDS-PAGE analysis, GST and GPx showed a single protein band of 25 and 18 kDa, respectively. They were able to retain their activities at an optimal temperature of 40°C for an hour and pH range 4-7. The 3D model of both enzymes was constructed showing helical structures, sheet and loops. Protein cavities were also detected to define druggable sites. GST model had two large pockets; 185Å3 and 71 Å3 with druggability score 0.5-0.8. For GPx, the pockets were relatively smaller, 71 Å3 and 32 Å3 with druggability score (0.65-0.66). Therefore, the present study showed that the consortium components as well as the stress-based conditions used could express both enzymes with enhanced productivity, recommending their application based on the obtained results.

Project description:BackgroundGlutathione (GSH) is one of the most important agents of the antioxidant defense system of the cell because, in conjunction with the enzymes glutathione peroxidase (GSH-Px) and glutathione S transferase pi (GSTpi), it plays a central role in the detoxification and biotransformation of chemotherapeutic drugs. This study evaluated the expression of GSH and the GSH-Px and GSTpi enzymes by immunohistochemistry in 30 canine mammary tumors, relating the clinicopathological parameters, clinical outcome and survival of the bitches. In an in vitro study, the expression of the genes glutamate cysteine ligase (GCLC) and glutathione synthetase (GSS) that synthesize GSH and GSH-Px gene were verified by qPCR and subjected to treatment with doxorubicin, to check the resistance of cancer cells to chemotherapy.ResultsThe immunohistochemical expression of GSH, GSH-Px and GSTpi was compared with the clinical and pathological characteristics and the clinical outcome in the bitches, including metastasis and death.The results showed that high immunoexpression of GSH was correlated to the absence of tumor ulceration and was present in dogs without metastasis (P < 0.05). There was significant correlation of survival with the increase of GSH (P < 0.05). The expression of the GSH-Px and GSTpi enzymes showed no statistically significant correlation with the analyzed variables (p > 0.05). The analysis of the relative expression of genes responsible for the synthesis of GSH (GCLC and GSS) and GSH-Px by quantitative PCR was done with cultured cells of 10 tumor fragments from dogs with mammary tumors.The culture cells showed a decrease in GCLC and GSS expression when compared with no treated cells (P < 0.05). High GSH immunoexpression was associated with better clinical outcomes.ConclusionTherefore, high expression of the GSH seems to play an important role in the clinical outcome of patients with mammary tumors and suggest its use as prognostic marker. The in vitro doxorubicin treatment significantly reduces the expression of GCLC and GSS genes so we can consider them to be candidates for predictive markers of therapeutic response in mammary cancer.

Dataset Information

Molecular Characterization and Gene Expression of Glutathione Peroxidase 1 in Tor tambroides Exposed to Temperature Stress.

Publications

Molecular Characterization and Gene Expression of Glutathione Peroxidase 1 in <i>Tor tambroides</i> Exposed to Temperature Stress.

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