• Title/Summary/Keyword: $P_{tac}$

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Overproduction and Operator DNA-Protein Blotting of R100 Mutant MerR from Shigella flexneri

  • Yoon, Kyung-Pyo
    • Journal of Microbiology and Biotechnology
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    • v.4 no.4
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    • pp.250-255
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    • 1994
  • Wild-type and four mutant R100 merR genes were cloned and the proteins overproduced under tac promoter control of pKK223-3. His118Ala, Cys117Ser, Cys126Ser, and wild-type MerR were successfully overproduced although amino-terminal 14 amino acids deletion mutant MerR was not successful. The amount of overproduced wild-type MerR protein as well as other mutant MerR was between 15%-20% of the total protein. The protein was able to be purified up to 95% homogeneity. Specific DNA-protein blotting experiments showed that the 95 bp operator containing DNA fragment could bind to Cys126Ser, His118Ala, and wild- type MerR, but not to Cys117Ser. These results were consistent with the previously reported complementation experiment results that His118Ala, Cys126Ser, and wild-type MerR could repress the mer operon but Cys117Ser could not.

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Expression of Yeast Antioxidant Protein Gene in E. coil (대장균(JM 109)에서 효모 Thiol-Specific Antioxident 단백질의 발현)

  • Kim, Il-Han
    • The Journal of Natural Sciences
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    • v.4
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    • pp.1-10
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    • 1991
  • The expression of the cloned Saccharomyces cerevisiae antioxidant protein gene in E. coil on plasmid was studied. Introduction of the DNA encoding yeast thiol-specific antioxidant protein into expression vector, PKK 223-3, having the tac promoter resulted in the production in E. coil of a Mr 27,000 polypeptide. This protein represented as much as about 1% of the bacterial soluble protein and showed the thiol-specific antioxidant properties. The bacterial protein showed physico-chemical properties indistringuishable from those of the yeast protein.

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Investigation of the Relationship between Protein, Message and Inducer Concentrations in Recombinant E. coli Cells

  • Jorgensen, Lene;Connor J. Thomas;Brian K. Oneill;Anton P.J. Middelbeg
    • Journal of Microbiology and Biotechnology
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    • v.7 no.1
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    • pp.21-24
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    • 1997
  • Chloramphenicol acetyl transferase (CAT) protein and mRNA levels in E. coli were determined following induction of a tac::cat construct by isopropyl-${\beta}$-thiogalactopyranoside (IPTG). High cat mRNA levels did not directly reflect CAT protein levels, in either shakeflask experiments or fermentations. Furthermore, concentrations of IPTG resulting in the highest levels of expression of cat mRNA, were different to those resulting in highest levels of CAT protein. The data suggest that high transcriptional activities lead to limitations at the translational level.

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Effect of Dietary Sodium Nitrate Consumption on Egg Production, Egg Quality Characteristics and Some Blood Indices in Native Hens of West Azarbaijan Province

  • Safary, H.;Daneshyar, Mohsen
    • Asian-Australasian Journal of Animal Sciences
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    • v.25 no.11
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    • pp.1611-1616
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    • 2012
  • The aim of the study was to evaluate the effect of sodium nitrate consumption on egg quality and quantity, and some blood parameters of native breeder hens of West Azerbaijan province. One hundred native hens were used from wk 25 to 32 of age. These birds were divided into two groups. One group was fed the control diet (CD) but the other fed the same diet supplemented with 4.2 g/kg sodium nitrate (ND). After 2 wks of adaptation, eggs were collected daily and egg mass and egg production were measured weekly for five weeks. To assess the egg quality parameters, two eggs from each replicate pen were collected for three consecutive days each week. At the end of experimental period (wk 32 of age), blood samples of 5 birds per replicate were collected from the wing vein into anticoagulant tubes. Dietary sodium nitrate didn't affect the egg production, shell stiffness, shell thickness and Haugh unit (p>0.05) but it decreased the both egg production and egg mass during the last three weeks (wks 30, 31 and 32) (p<0.05). Furthermore, a treatment effect was observed for yolk colour (p<0.05). Both the egg production and egg mass were increased over time (p<0.05). No significant treatment${\times}$time interaction was observed for egg weight, egg production and egg mass (p>0.05). No effect of time or treatment${\times}$time were observed for shell stiffness (p>0.05). Over time, shell thickness was decreased while Haugh unit increased (p<0.05). None of the blood TP and TG or the activity of ALT, AST and LDH enzymes were affected by dietary consumption of sodium nitrate at wk 32 of age (p>0.05). Sodium nitrite decreased both the TAC and TC at wk 32 of age (p<0.001). It was concluded that the lower body antioxidant capacity of nitrate fed birds resulted in the lower performance (egg weight, egg production and egg mass).

Allium Jesdianum Extract Improve AcetaminophenInduced Hepatic Failure through Inhibition of Oxidative/Nitrosative Stress

  • Sohrabinezhad, Zohreh;Dastan, Dara;Asl, Sara Soleimani;Nili-Ahmadabadi, Amir
    • Journal of Pharmacopuncture
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    • v.22 no.4
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    • pp.239-247
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    • 2019
  • Objectives: Allium jesdianum (Aj) is a medicinal plant that has highlighted pharmacological features. In this study, the effects of Aj extract were examined on acetaminophen (APAP)-induced hepatic failure in rats. Methods: Methanolic fraction of hydro-alcoholic extract of Aj was obtained by silica gel column chromatography method. Animals were randomly divided into four groups each containing six rats and treated by gavage as follows: the first and second groups received normal saline, the third and fourth groups were received with 50 and 100 mg/kg of Aj extract, respectively. After two consecutive weeks, the groups 2-4 were given a single dose of APAP (2 g/kg). After 48 hours, blood and liver samples were collected for biochemical and histological examinations. Results: The findings of the study demonstrated that APAP caused a significant increase in ALT (P < 0.001), AST (P < 0.001), LDH (P < 0.001), ALP (P < 0.001) serum levels, hepatic lipid peroxidation (LPO; P < 0.001) and nitric oxide (NO; P < 0.001). In this regard, APAP led to the depletion of the total antioxidant capacity (TAC; P < 0.001), glutathione and total thiol groups (TTGs; P < 0.001), and structural change in the liver. In the Aj extract groups, a considerable improvement was found in the hepatic function alongside the histopathologic changes. Conclusion: This investigation indicated that the influential effects of Aj extract in APAP-induced hepatic failure might depend on its effect on improving oxidant/antioxidant balance in hepatic tissue.

High-Level Expression of Human Cytochrome P450 3A4 by Co-Expression with Human Molecular Chaperone HDJ-1 (Hsp40)

  • Ahn, Tae-Ho;Yun, Chul-Ho
    • Archives of Pharmacal Research
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    • v.27 no.3
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    • pp.319-323
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    • 2004
  • Cytochrome P450 (CYP) 3A4 is of great interest because of its important roles in the oxidation of numerous drugs and xenobiotics. HDJ-1, a molecular chaperone in human, is known to assist the correct folding of unfolded proteins. To achieve a high yield of recombinant human CYP3A4 in Escherichia coli, the CYP3A4 encoding gene was co-expressed with the chaperone HDJ-1, under the control of an inducible tac promoter in a bicistronic format. The levels of expression of the CYP3A4 in the bicistronic construct reached up to 715 nmol $(liter culture)^{-1}$ within 16 h at $37^{\circ}C$, which was about a 3.3-fold increase compared to that of the CYP3A4 alone without the HDJ-1. By co-expression with HDJ-1, the catalytic activity of CYP3A4 was also increased by -15-fold. The amount of activity increase was similar to that of the CYP production at the whole cell level. The present over-expression system may be useful for the rapid production of large amounts of active CYP3A4 in E. coli.

Design and Cloning of the Gene for a Novel Insulin Analogue, $(B^{30}$-Homoserine) Human Insulin

  • Nam, Doo-H.;Ko, Jeong-Heon;Lee, Seung-Yup
    • Archives of Pharmacal Research
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    • v.16 no.4
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    • pp.271-275
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    • 1993
  • In order to prepare a novel human insulin analogue suhbstituted with homoserine at B$^{30}$ / position, (B$^{30}$ /-homoserine) human insulin, a synthetic gene was designed by linking directly a gene for B chain with that for A chain. This gene was constructed by enzymatic joining of 10 different synthetic oligonucleotides, and then inserted at the polylinker region of pUC19 plasmid. To achieve a high level of gene expression, the gene fusion technique region of pUC19 plasmid. To achieve a high level of gene expression, the gene fusion technique was employed using amino terminal regions of lacZ gene up to Clal or hpal, and either of them has been located under tac promoter. The chemical induction of these fused genes by isopropyl-.betha.-D-thiogalactopyranoside (IPTG) gave a satisfactory level of expression in Escherichia coli harboring the ocnstructed plasmids. It was observed that the fused gene product as a single chain insulin precusor was produced more than 30% of total cell protein of E. coli as a form of inclusion body.

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Effect of Broussonetia papyrifera L. silage on blood biochemical parameters, growth performance, meat amino acids and fatty acids compositions in beef cattle

  • Tao, Hui;Si, Bingwen;Xu, Wencai;Tu, Yan;Diao, Qiyu
    • Asian-Australasian Journal of Animal Sciences
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    • v.33 no.5
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    • pp.732-741
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    • 2020
  • Objective: The study was conducted to investigate the effects of Broussonetia papyrifera L. (B. papyrifera) silage on growth performance, serum biochemical parameters, meat quality, and meat amino acids and fatty acids compositions in beef cattle. Methods: Sixty-four male Angus beef cattle were assigned to 4 groups with 4 pens in each group and 4 beef cattle in each pen, and fed with the total mixed ration supplemented with 0%, 5%, 10%, or 15% B. papyrifera silage for 100 days (control group, 5% group, 10% group and 15% group) separately. Results: Beef cattle had significantly higher final body weight (BW) in 15% group, higher average daily gain (ADG) and dry matter intake (DMI) in 5% group, 10% group and 15% group, and higher feed conversion ratio (FCR) in 10% group and 15% group. Significantly higher blood superoxide dismutase (SOD) concentration was noted in 15% group, higher blood total antioxidant capacity (TAC) in 10% group and 15% group, lower 8-hydroxydeoxyguanosine (8-OHdG) and malondialdehyde (MDA) in 15% group. Meat had lower pH in 15% group, higher Commission International DeI'Eclairage (CIE) L in 5% group, 10% group, and 15% group, and lower drip loss in 15% group. Greater concentration of meat polyunsaturated fatty acids (PUFA) was observed in 10% group and 15% group, and docosahexaenoic acid (DHA) in 15% group. Conclusion: Diet with 15% B. papyrifera silage could improve performance and increase final BW, ADG, DMI, and FCR, enhance the antioxidant functions by decreasing blood 8-OHdG and MDA and increasing blood SOD and TAC, improve the meat quality by lowing pH and drip loss and increasing CIE L, increase the meat PUFA and DHA concentration. Polyphenols and flavonoids might be the main components responsible for the antioxidant activity and anti-biohydrogenation in the B. papyrifera silage. And B. papyrifera silage could be used as a new feedstuff in beef cattle nutrition.

Plasma adipocytokines and antioxidants-status in Korean overweight and obese females with dyslipidemia

  • Ha, Ae Wha;Jeong, Su Youn;Kang, Nam E;Kim, Woo Kyoung
    • Nutrition Research and Practice
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    • v.8 no.4
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    • pp.417-424
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    • 2014
  • BACKGROUD/OBEJECTIVES: It is hypothesized that obese people with dyslipidemia is more likely to have increased oxidative stress and decreased antioxidant status, in comparison with the controls who were obese without dyslipidemia. Thus, the aims of the present study were to determine the dietary intakes, plasma adipokines, and antioxidative systems between obese with dyslipidemia and obese without dyslipidemia were investigated. SUBJECTS/METHODS: Female subjects who were between 20 and 55 years old, and whose BMI was 23 or greater were recruited. Subjects who met the criteria of $BMI{\geq}23$, total cholestero ${\geq}200mg/dL$, LDL cholesterol ${\geq}130mg/dL$, and $TG{\geq}110mg/dL$ were categorized Obese with dyslipidemia. Anthropometric measurements and blood biochemical tests were conducted. The diet survey was conducted by a trained dietitian using two days of 24 hour dietary recall. The lipid peroxidation, the plasma total antioxidant capacity (TAC), the activities of antioxidantive enzymes, and various antioxidantive vitamins levels were determined. RESULTS: Plasma adiponectin and leptin levels were also determined. There were no significant differences for age, Body Mass index (BMI), and body fat (%), waist-size between two groups. Obese with dyslipidemia had significantly high levels of total cholesterol, triglyceride, LDL-cholesterol, the ratio of total cholesterol/HDL-C, and the ratio of HDL-C/LDL-C, respectively. Blood alkaline phosphatase level was statistically different between the two groups (P < 0.05). No statistical significance in dietary intake between two groups was shown. In case of obese with dyslipidemia group, the levels of GSH-Px (P < 0.05) and catalase (P < 0.05) as well as adjusted blood retinol (P < 0.05) and tocopherol level (P < 0.05) were significantly low. However, the plasma concentration of leptin was significantly high (P < 0.05). CONCLUSIONS: Obesity with dyslipidemia was shown to have high arthtrogenic index, depleted antioxidant status, and higher blood leptin levels which suggest higher risks of oxidative stress and cardiovascular diseases.