• Journal of Nutrition and Health
• /
• 제48권6호
• /
• pp.468-475
• /
• 2015

우리나라 일부 건강한 유아를 대상으로 Mn-SOD Val16Ala, GSTP1 Ile105Val, GSTT1 present/null, GSTM1 present/null 유전자 다형성 분포를 살펴본 결과, Mn-SOD Val/Val형, GSTP1 Ile/Ile형, GSTT1 null 형, GSTM1 null 형이 주된 (major) 유전자형인 것으로 나타났다. 이 중 Mn-SOD Val/Val형은 Val/Ala 또는 Ala/Ala형에 비해 소변 8-OHdG 수준이 유의적이지는 않으나 높은 경향을 나타내었고, GSTP1 Ile/Ile형은 Ile/Val 또는 Val/Val형에 비해 소변 8-OHdG 수준이 유의적으로 낮았다. 간접흡연에의 노출 여부와 간접흡연-유전자 다형성의 상호 작용이 산화손상지표에는 유의적인 영향을 미치지 않는 것으로 나타났다. 이상의 결과로 볼 때 건강한 유아에서 GSTP1 Val allele 보유한 경우 산화적 손상에 대해 취약할 수 있음을 제시하며, 추후 대규모 연구를 통한 검증 및 이들 유전자형을 보유한 대상자를 위한 효과적인 영양 중재방안에 대한 고려가 필요할 것으로 사료된다.

• 한국미생물·생명공학회지
• /
• 제25권2호
• /
• pp.137-143
• /
• 1997

Porcine transforming growth factor-$\beta$1 (TGF-$\beta$1) was expressed in Escherichia coli using cDNA of TGF-$\beta$1 and glutathione S-transferase (GST) fusion vector pGEX-1$\lambda$T. An ApoI-Tth111I fragment of cDNA which correspond to the amino acid residues from 123 to 390 of the precursor TGF-$\beta$1 was inserted into EcoRI-Tth111I digested pGEM#-l$\lambda$T and the recombined plasmid was named pGET-12. Gene products from the cloned regions of the recombinant plasmids pGET-12 was not detected in soluble fraction of cell free extract but detected in insoluble fraction. The solubilization of insoluble gene product was achieved by the treatment of N-laurylsarcosine. Molecular weight of partially purified proteins determined by electrophoresis was same as expected from cloned fragment. The ELISA test results of the purified proteins showed that immunologically detectable epitope was preserved in recombinant protein.

• 대한예방한의학회지
• /
• 제3권2호
• /
• pp.183-210
• /
• 1999

In order to investigate the protective effect of Injinhotanghapsihosogantang-gagambang on the liver injury of rats induced by $CCl_4$ and d-galactosamine, the serum transaminase(GOT&GPT) alkaline phosphatase(ALP), lactic dehydrogenase(LDH) for enzyme activities, triglyceride for serum component, liver weight and glutathione S-transferase(GST), Superoxide dimutase(SOD) were measured. All animals were divided into 5 groups, those were normal group(untreated), control group(treated with vehicle 0.9% Saline solution), sample I group(10mg/kg administrated), sample II group(30mg/kg administrated), Silymarin 200 administrated group. The results were as follows: 1. The inhibitory effects of the serum GOT activities in rats induced by $CCl_4$ were noted in both sample I (p<0.001) and sample II group(p<0.001). In serum GPT activities, sample I (p<0.01) and sample II group(p<0.01). 2. The inhibitory effects of the serum LDH activities in rats induced by $CCl_4$ were noted in both sample I (p<0.001) and sample II group(p<0.001). 3. The increased effects of the serum ALP activities in rats induced by $CCl_4$ were not recognized. 4. The inhibitory effects of the serum triglyceride content level in rats induced by $CCl_4$ were inhibited in only sample II group(p<0.05). 5. The increased effects of the liver weight level in rats induced by $CCl_4$ were inhibited in both sample I (p<0.05) and sample II group(p<0.05). 6. The inhibitory effects of the GST activities in rats induced by $CCl_4$ were not recognized. In SOD activities, both sample I (p<0.05) and sample II group(p<0.001) showed the inthbitory effects. 7. The inhibitory effects of in the serum GOT, GPT activities in rats induced by d-galactosamine were not recognized. 8. The increases of the serum LDH level in rats induced by d-galactosamine were noted in both sample I (p<0.01) and sample II group(p<0.001). 9. The inhibitory of the serum triglyceride content level in rats induced d-galactosamine were noted in only sample II group(p<0.05). According to the above results, it is considered that Injinhotanghapsihosogantang-gagambang has protective effect against liver injury in rats induced by $CCl_4$ and d-galactosamine.

• Molecules and Cells
• /
• 제37권3호
• /
• pp.264-269
• /
• 2014

The molecular interaction between tumor suppressor p53 and the anti-apoptotic Bcl-2 family proteins plays an essential role in the transcription-independent apoptotic pathway of p53. In this study, we investigated the binding of p53 DNA-binding domain (p53DBD) with the anti-apoptotic Bcl-2 family proteins, Bcl-w, Mcl-1, and Bcl-2, using GST pull-down assay and NMR spectroscopy. The GST pull-down assays and NMR experiments demonstrated the direct binding of the p53DBD with Bcl-w, Mcl-1, and Bcl-2. Further, NMR chemical shift perturbation data showed that Bcl-w and Mcl-1 bind to the positively charged DNA-binding surface of p53DBD. Noticeably, the refined structural models of the complexes between p53DBD and Bcl-w, Mcl-1, and Bcl-2 showed that the binding mode of p53DBD is highly conserved among the anti-apoptotic Bcl-2 family proteins. Furthermore, the chemical shift perturbations on Bcl-w, Mcl-1, and Bcl-2 induced by p53DBD binding occurred not only at the p53DBD-binding acidic region but also at the BH3 peptide-binding pocket, which suggests an allosteric conformational change similar to that observed in Bcl-$X_L$. Taken altogether, our results revealed a structural basis for a conserved binding mechanism between p53DBD and the anti-apoptotic Bcl-2 family proteins, which shed light on to the molecular understanding of the transcription-independent apoptosis pathway of p53.

• Biotechnology and Bioprocess Engineering:BBE
• /
• 제7권3호
• /
• pp.155-162
• /
• 2002

The cyclic amidohydrolase family enzymes, which include allantoinase, dihydroorotase, dihydropyrimidinase and (phenyl)hydantoinase, are metal-dependent hydrolases and play a crucial role in the metabolism of purine and pyrimidine in vivo. Each enzyme has been independently characterized, and thus well documented, but studies on the higher structural traits shared by members of this enzyme family are rare due to the lack of comparative study. Here, we report upon the expression in E. coli cells of maltose-binding protein (MBP)- and glutathione S-transferase (GST)-fused cyclic amidohydrolase family enzymes, facilitating also for both simple purification and high-level expression. Interestingly, the native quaternary structure of each enzyme was maintained even when fused with MBP and GST. We also found that in fusion proteins the favorable biochemical properties of family enzymes such as, their optimal pHs, specific activities and kinetic properties were conserved compared to the native enzymes. In addition, MBP-fused enzymes showed remarkable folding ability in-vitro. Our findings, therefore, suggest that a previously unrecognized trait of this family, namely the ability to functional fusion with some other protein but yet to retain innate properties, is conserved. We described here the structural and evolutionary implications of the properties in this family enzyme.

• Journal of Plant Biology
• /
• 제37권2호
• /
• pp.141-149
• /
• 1994

Levels of genetic diversity, population genetic structure, and gene flow in Hosta yingeri, a herbaceous perennial endemic to Taehuksan, Sohuksan, and Hong Islands, were investigated. Starch gel electrophoresis was conducted on leaves for 101 plants collected from three populations. Although the distribution of thespecies is restricted in the islands, it maintains high levels of genetic variatin; 64% of polymorphic loci in at least one population (Ps), the mean number of alleles per locus (Ap) of 1.92, and the mean effective number of alleles per locus (Aep) of 1.52. Overall, mean genetic diversity (Hep=0.250) was substantially higher than mean estimate for species with very similarlife history traits (0.102). Large populaton size, the persistence of multiple generations within populations, high fecundity, predominantly outcrossing breeding system, large size of pollinator visitation areas may be explanatory factors contributing the higher level of genetic diversity maintained within populations. Analysis of fixation indices showed an overall slight excess of heterozygotes (mean FIS=-0.066) relative to Hardy-Weinberg expectations, which may in part be due to the near self-incompatible breeding system in the species. Significant differences in allele frequencies among populaitns were found for 14 out of 16 polymorphic loci (P<0.05). Slightly more than 80% of the total variation in the species was common to all populations (GST=0.198). As expected, indirect estimate of the number of migrants per generation (Nm=0.45, calculated from mean GST) and nine private alleles found in the three populations indicate that gene movement among three isolated island populations was low.

• Journal of Nutrition and Health
• /
• 제29권10호
• /
• pp.1080-1086
• /
• 1996

The purpose of this study was to determine the effects of taurine supplementation on the hepatic lipid peroxidation, activiteis of defense enzymes and membrane stability during rat hepatocarcinogenesis. Hepatocarcinogenesis was induced by Solt & Farber modification. Lipid peroxide contents of carcinogen treated group which was not supplemented with taurine were lower than those of control group. This might be that peroxide is decreased because of the activation of detoxifing enzyme. Glutathione S-transferase(GST) activites of carcinogen treated groups were significantly (p<0.05) increased compared to those of control groups. The GST activities of group supplemented with taurine before treatment of carcinogen and during the all period of experiment were only less increased. In carcinogen treated groups, glutathione peroxidase(GPx) activites of groups supplemented with taurine were higher than those of non supplemented group. By carcinogen treatemtn, glucose 6-phosphatase(G6Pase) activites, index of membrane stability were decreased, but in carcinogen treated groups supplemented with taurine, they were less decreased. These results suggest that taurine supplementation seems to inhibit lipid peroxidation, to change the activities of defense enzymes and to prevent to membrane disintegration during chemically induced hepatocarcinogenesis.

• 대한한의학방제학회지
• /
• 제11권2호
• /
• pp.135-146
• /
• 2003

Objectives : Wolguk-whan has been used as a prescription of natural drug for the treatment of stress digestive system disease. Recently, we reported that Wolguk-whan methnol extract (WGWM) exerted a significant protective effect against oxidative damage to the liver of ICR mice. This study was purposed to investigate the effects of Wolguk-whan water extract (WGWW) on liver injury induced by oxidative stress. Methods : In order to investigate the effects of WGWW on acute liver injury, ICR mice were pretreated with WGWW for 6days, starved for 24hrs, and administerated acetamirtophen(500mg/kg, i.p.). In the liver homogenates, lipid peroxide and glutathione(GSH) levels were measured. In addition, activities of hepatic enzyme, such as catalase, glutathione peroxidase(GSH-Px), glutathione S-transferase(GST) were measured in the hepatic mitochondrial and cytosolic fractions. Results : In vivo administeration of WGWW showed effective inhibition of acetaminophen induced lipid peroxidation, and showed elevations of GSH level, catalase, GSH-Px, GST activities. Conclusions : These results suggested that WGWW might suppress the formation of oxidative metabolites, and prevent acetaminophen induced hepatotoxicity.

• Journal of Microbiology and Biotechnology
• /
• 제19권12호
• /
• pp.1620-1627
• /
• 2009

The design and expression of an antihypertensive peptide multimer (AHPM), a common precursor of 11 kinds of antihypertensive peptides (AHPs) tandemly linked up according to the restriction sites of gastrointestinal proteases, were explored. The DNA fragment encoding the AHPM was chemically synthesized and cloned into expression vector pGEX-3X. After an optimum induction with IPTG, the recombinant AHPM fused with glutathione S-transferase (GST-AHPM) was expressed mostly as inclusion body in Escherichia coli BL21 and reached the maximal production, 35% of total intracellular protein. The inclusion body was washed, dissolved, and purified by cation-exchange chromatography under denaturing conditions, followed by refolding together with size-exclusion chromatography and gradual dialysis. The resulting yield of the soluble GSTAHPM (34 kDa) with a purity of 95% reached 399 mg/l culture. The release of high active fragments from the AHPM was confirmed by the simulated gastrointestinal digestion. The results suggest that the design strategy and production method of the AHPM will be useful to obtain a large quantity of recombinant AHPs at a low cost.

• Journal of Microbiology and Biotechnology
• /
• 제13권5호
• /
• pp.738-744
• /
• 2003

The gene ADH encoding NAD-dependent alcohol dehydrogenase from Bacillus stearothennophilus was cloned and overexpressed as a GST fusion protein at a high level in Escherichia coli. The expressed fusion protein was purified simply by glutathione affinity chromatography. GST fusion protein was then cleaved by thrombin, while soluble enzyme was further purified by glutathione affinity chromatography. The recombinant enzyme had the same elctrophoretic mobility as the native enzyme from Bacillus stearothennophilus. The recombinant enzyme catalyzed the oxidation of a number of alcohols and exhibited high activities towards secondary alcohols. The $K_m\;and\;V_{max}$ values of the recombinant enzyme for ethanol were 5.11 mM and 61.35 U/mg, respectively. Pyridine and imidazole notably inhibited the enzymatic activity. The activity of the recombinant enzyme optimally proceeded at pH 9.0 and $70^{\circ}C$. The midpoint of the temperature-stability curve for the recombinant enzyme was approximately $68^{\circ}C$, and the enzyme was not completely inactivated even at $85^{\circ}C$. The recombinant enzyme showed a high resistance towards denaturing agents (0.05% SDS, 0.1 M urea). Therefore, due to its stability and relatively broad substrate specificity, the recombinant enzyme could be utilized in bio-industrial processes and biosensors.