1.Induction of Anticarcinogenic Enzymes of Waxy Brown Rice Cultured with Phellinus igniarius 26005.
Ki Bum PARK ; Hyo Cheol HA ; So Yeun KIM ; Hyo Jeong KIM ; Jae Sung LEE
Mycobiology 2002;30(4):213-218
The induction of NAD(P)H: quinone oxidoreductase (QR), glutathione S-transferase (GST), and glutathione (GSH) levels in hepa1c1c7 cells (murine hepatoma) by waxy brown rice cultured with Phellinus igniarius to induce anticarcinogenic enzymes were measured. In addition, the inhibition of polyamines metabolism was tested with the growth of Acanthamoeba castellanii. The result shows that QR, GST activities, and GSH levels of experimental animals were increased much more by feeding the methanol extract of waxy brown rice cultured with Phellinus igniarius than those of the rats received the ethanol of uncultured brown rice. The growth of A. castellanii was inhibited mostly at 40 mg/3 ml concentration of methanol extract of waxy brown rice cultured with P. igniarius. The results suggested that waxy brown rice cultured with P. igniarius possess chemopreventive activity by inducing anticarcinogenic enzymes and inhibiting polyamine metabolism.
Acanthamoeba castellanii
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Animals
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Chemoprevention
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Ethanol
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Glutathione
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Glutathione Transferase
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Metabolism
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Methanol
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Polyamines
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Rats
2.Effect of trans-acting factor on rat glutathione S-transferase P1 gene transcription regulation in tumor cells.
Dongyuan LIU ; Mingxiang LIAO ; Jin ZUO ; Fude FANG
Chinese Medical Journal 2002;115(1):103-106
OBJECTIVETo investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.
METHODSThe binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer I (GPEI) and glutathione S-transferase P enhancer II-1 (GPE II-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment.
RESULTSTrans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPE II-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver.
CONCLUSIONcGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.
Animals ; Carrier Proteins ; metabolism ; Enhancer Elements, Genetic ; physiology ; Gene Expression Regulation, Enzymologic ; Glutathione S-Transferase pi ; Glutathione Transferase ; genetics ; Isoenzymes ; genetics ; Nuclear Proteins ; metabolism ; Rats ; Transcription, Genetic
3.A study on the relationship between glutathione S-transferases gene polymorphism and susceptibility response to hypoxia.
Hui-qin YAN ; Xue-chuan SUN ; Kong-xiang LIU ; Sheng-wei WANG ; Tao LIU
Chinese Journal of Applied Physiology 2006;22(3):334-337
AIMTo investigate the relationship between glutathione S-transferases gene polymorphism and susceptibility response to hypoxia.
METHODSIn the case-control study, the gene polymorphisms of glutathione S-transferases were tested in Tibetan mountaineers and sea-level Han Chinese by multiple-PCR and PCR-RELP.
RESULTSThe frequency of GSTT1 null genotype was significant different between Tibetan mountaineers and sea-level Han Chinese (P < 0.05), OR = 1.86 (95% CI = 1.01-3.39), and also for GSTP(1-105) mutant genotype in two groups (P < 0.01), OR = 2.19 (95% CI = 1.16-4.13). There was significant difference between A allele and G allele of GSTP(1-105) groups (P < 0.01). There was no difference for GSTM1 null genotype between two groups (P > 0.05), OR = 0.78 (95% CI = 0.43 - 1.42).
CONCLUSIONGSTT1 and GSTP(1-105) genotype may be associated with susceptibility response to altitude hypoxia.
Adult ; Alleles ; China ; Genotype ; Glutathione S-Transferase pi ; genetics ; Glutathione Transferase ; genetics ; Humans ; Hypoxia ; genetics ; Male ; Mountaineering ; Polymorphism, Genetic ; Reactive Oxygen Species ; metabolism ; Young Adult
4.Advances in plant anthocyanin transport mechanism.
Lu WANG ; Silan DAI ; Xuehua JIN ; He HUANG ; Yan HONG
Chinese Journal of Biotechnology 2014;30(6):848-863
Anthocyanin biosynthesis is one of the thoroughly studied enzymatic pathways in biology, but little is known about the molecular mechanisms of its final stage: the transport of the anthocyanins into the vacuole. A clear picture of the dynamic trafficking of flavonoids is only now beginning to emerge. So far four different models have been proposed to explain the transport of anthocyanins from biosynthetic sites to the central vacuole, and four types of transporters have been found associated with the transport of anthocyanins: glutathione S-transferase, multidrug resistance-associated protein, multidrug and toxic compound extrusion, bilitranslocase-homologue. The functions of these proteins and related genes have also been studied. Although different models have been proposed, cellular and subcellular information is still lacking for reconciliation of different lines of evidence in various anthocyanin sequestration studies. According to the information available, through sequence analysis, gene expression analysis, subcellular positioning and complementation experiments, the function and location of these transporters can be explored, and the anthocyanin transport mechanism can be better understood.
Anthocyanins
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metabolism
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Biological Transport
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Glutathione Transferase
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metabolism
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Membrane Transport Proteins
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metabolism
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Multidrug Resistance-Associated Proteins
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metabolism
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Plants
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metabolism
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Vacuoles
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metabolism
6.Large recombinant protein displayed on filamentous phage surface and its interaction with small molecule.
Bo KONG ; Hui LIU ; Sheng-Li YANG ; Wei-Jun MA
Chinese Journal of Biotechnology 2006;22(1):19-25
Recombinant proteins were expressed as fusions with the phagemid system of pHEN-KM13 and the characteristics and activities of the fusion proteins displayed on the surface of filamentous phagintain the were studiedon abilty. The altered titer of rescued phages from the phagemid system after trypsin treatment indicated the relative quantity of the phages displaying fusion proteins. The rescue phages displayed foreign proteins could keep the bacterial infection ability, while the bald phage without foreign protein displayed on its surface was sensitive to trypsin treatment and lost the bacterial infection ability. To determine the upper limit for filamentous phage display, four recombinant proteins, glutathione-S-transferase and glutathione-S-transferase fused with three various size peptide linkers, were fused to N terminus of capsid protein gp3 and rescued by helper phage KM13. The rescued phages which displayed fused protein with the size of 40kD or less maintain the infection ability. To assay the activity of the phage displayed protein, the known small molecule probe was used in the interaction study with protein incorporated on phage surface. Results showed that the glutathione-S-transferase on phage surface still bound to glutathione specifically. It indicated that the glutathione-S-transferase displayed on phage surface was correctly folded and functionally active. The results demonstrated that it was feasible to use small molecule probes to interact with the protein displayed on phage surface. In turn, the method described here also demonstrated that phage display system could be utilized to investigate the interactions between protein and small molecules.
Glutathione
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genetics
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metabolism
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Glutathione Transferase
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biosynthesis
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genetics
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Helper Viruses
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genetics
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metabolism
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Inovirus
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genetics
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metabolism
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Peptide Library
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Protein Binding
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Protein Interaction Domains and Motifs
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genetics
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Recombinant Fusion Proteins
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biosynthesis
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genetics
7.Effect of modified ricin on the reduction of hepatotoxicity in mice.
Wen-li LI ; Chun-xu HAI ; Ying ZHAO ; Xin LIANG ; Wen-xue WANG
Chinese Journal of Industrial Hygiene and Occupational Diseases 2003;21(3):209-211
OBJECTIVETo observe the significance of ricin (RT) with chemical nidification to reduce the hepatotoxicity in mice and its anticancer effect.
METHODSMice were exposed to RT and RT-PDP [ricin chemically modified by N-succinimidyl 3-(2-pyridyldithio)-propionate, (SPDP)] respectively, and their serum activity of glutathione-S-transferase (GST) and liver glutathione (GSH) content were determined. The ultramicro-structure under electron microscope was also observed.
RESULTSThe GST activity increased with doses, and the increase in ricin group was higher than that in RT-PDP group; the activities of GST in RT group at 12.5, 15.0 micro g/kg [(93.65 +/- 12.30), (153.71 +/- 26.64) IU/L respectively] were higher than those in RT-PDP group [(62.97 +/- 11.22), (78.20 +/- 15.71) IU/L] (P < 0.05). The contents of GSH were decreased with doses; but the contents of GSH in RT-PDP group at 2.5, 5.0, 7.5, 10.0, 15.0 micro g/kg [(6.34 +/- 1.43), (4.14 +/- 1.82), (3.54 +/- 0.64), (2.73 +/- 1.82), (1.82 +/- 0.62) micro mol/L respectively] were still higher than those in RT group [(3.53 +/- 0.95), (2.12 +/- 0.54), (1.82 +/- 0.71), (1.52 +/- 0.34), (0.81 +/- 0.36) micro mol/L] (P < 0.01). Electron microscopic examination showed that the injury of liver cells in RT group was more severe than that in RT-PDP group.
CONCLUSIONThe hepatotoxicity of ricin in mice may be reduced by chemical modification.
Animals ; Chemical Warfare Agents ; toxicity ; Female ; Glutathione ; metabolism ; Glutathione Transferase ; metabolism ; Liver ; drug effects ; metabolism ; ultrastructure ; Male ; Mice ; Microscopy, Electron ; Ricin ; analogs & derivatives ; toxicity
8.Molecular characteristics of two Phi glutathione S-transferases in Selaginella moellendorffii.
Yuanjie ZHANG ; Zhiling YANG ; Hailing YANG
Chinese Journal of Biotechnology 2016;32(7):927-936
Glutathione S-transferase (GST) is important in plants to resist various stresses. In this study, two Phi GST genes (SmGSTF1 and SmGSTF2) were cloned from Selaginella moellendorffii. SmGSTF1 and SmGSTF2 genes encode proteins of 215 amino acid residues. Gene expression analysis showed that the two genes were expressed in roots, stems and leaves. The recombinant SmGSTF1 and SmGSTF2 proteins were overexpressed in Escherichia coli, and purified by Ni-affinity chromatography. SmGSTF1 and SmGSTF2 had the catalytic activity towards 1-Chloro-2,4-Dieitrobenzene, 4-Chloro-7-nitro-1,2,3-benzoxadiazole (NBD-Cl), and 4-Nitrobenzyl chloride substrates. SmGSTF1 also had the activity towards Fluorodifen and Cumyl hydroperoxide (Cum-OOH), whereas SmGSTF2 not. The enzyme kinetics analysis showed that SmGSTF1 and SmGSTF2 had high affinity towards glutathione, and low affinity towards 1-Chloro-2, 4-Dieitrobenzene. The enzymatic activity of SmGSTF1 and SmGSTF2 had high catalytic activity between pH 7 and 8.5, and between 45 and 55 °C. SmGSTF1 and SmGSTF2 may have an important role in the resistance of Selaginella moellendorfii against stress.
Amino Acid Sequence
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Cloning, Molecular
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Escherichia coli
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Glutathione Transferase
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genetics
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metabolism
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Plant Proteins
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genetics
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metabolism
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Selaginellaceae
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enzymology
9.Screening and identification interaction proteins of connexin 30.
Ding-hua HE ; Yong FENG ; Ling-yun MEI ; Chu-feng HE
Chinese Journal of Otorhinolaryngology Head and Neck Surgery 2009;44(9):758-761
OBJECTIVETo explore interaction proteins affect functions of connexin 30 (Cx30) by screening and identification interaction proteins of Cx30.
METHODSThe fusion expression vecto of CX30-C-terminal functional domain-pGEX-4T-2-GST was constructed, and then, fusion protein and GST were purified. They were incubated with the proteins of the foetus brain tissue disruption to pull down interaction proteins. The interaction proteins were separated by SDS-PAGE. Differential straps were cut to enzymolysis to prepare for mass chromatographic analysis, and then to index and screen interaction proteins in NCBInr database. The interaction proteins were identified by immunolocalization.
RESULTSThe four interaction proteins of Cx30 were screened in the foetus brain tissue, as follow, Keratin 16, Camk2b, Tubulin beta-3 and alpha-tubulin. Cx30 was proved to coexist with Keratin 16 and Tubulin beta-3.
CONCLUSIONSKeratin 16, Camk2b, Tubulin beta-3 and alpha-tubulin are the interaction proteins of Cx30. The interaction proteins affect the assembly, intracellular transport, and channel switch of Cx30.
Connexin 30 ; Connexins ; genetics ; metabolism ; Genetic Vectors ; Glutathione Transferase ; Humans ; Mutagenicity Tests ; Protein Interaction Mapping ; Recombinant Proteins ; genetics ; metabolism
10.Effect of microencapsulation on the expression of the oxidative stress genes of HepG2 cells and exogenous regulation.
Jing XIAO ; Ying ZHANG ; Weiting YU ; Wei WANG ; Xiaojun MA
Journal of Biomedical Engineering 2014;31(2):373-378
The aim of this research is to investigate the influence of microencapsulation on the expression of the oxidative stress genes and exogenous regulation of HepG2 cells. We compared the expression of hemeoxygenase-1 (HO-1) and glutathione S-transferases-A1 (GST-A1) in HepG2 cells under different culture conditions through real-time PCR. The effects of exogenous antioxidants on cell viability and albumin levels were also evaluated through MTT assay and ELISA assay. The results showed that after culturing for 6 and 16 days, the expression levels of HO-1 in encapsulated cells were approximately 4.9 and 3.1 times higher than that of monolayer cells at the same culture period; As for the expression levels of GST-A1, they were elevated to 11.2 and 33 times of monolayer cells (P < 0.05). Accordingly, we found that NAC at 5-10 mmol/L significantly increased the viability by 40%-70% and the biosynthetic function by 20%-30% in microencapsulated HepG2 cells (P < 0.05). GSH increased the viability of the encapsulated cells by 20%-55% and the biosynthetic function by 15% (P < 0.05). In conclusion, oxidative stress exists in the microcapsules and affects genes expression. Exogenous antioxidants can prevent the inhibition effects of oxidative stress on cellular growth.
Antioxidants
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pharmacology
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Cell Survival
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Gene Expression Regulation
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Glutathione Transferase
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metabolism
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Heme Oxygenase-1
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metabolism
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Hep G2 Cells
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Humans
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Oxidative Stress