• Journal of Life Science
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• v.6 no.3
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• pp.185-192
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• 1996

Bacteriophage T7 gene 2.5 protein, a single-stranded DNA binding protein, is required for T7 DNA replication, recombination, and repair. T7 gene 2.5 protein has two distinctive domains, DNA binding and C-terminal domain, directly involved in protein-protein interaction. Gene 2.5 protein participates in the DNA replication of Bacteriophage T7, which makes this protein essential for the T7 growth and DNA replication. What gene 2.5 protein makes important at T7 growth and DNA replication is its binding affinity to single-stranded DNA and the protein-protein important at T7 DNA replication proteins which are essential for the T7 DNA synthesis. We have constructed pGST2.5(WT) encoding the wild-type gene 2.5 protein and pGST2.5$\Delta $21C lacking C-terminal 21 amino acid residues. The purified GST-fusion proteins, GST2.5(WT) and GST2.5(WT)$\Delta$21C, were used for whether the carboxyl-terminal domain participates in the protein-protein interactions or not. GST2.5(WT) and GST2.5$\Delta$21C showed the difference in the protein-protein interaction. GST2.5(WT) interacted with T7 DNA polymerase and gene 4 protein, but GST2.5$\Delta$21C did not interact with either protein. Secondly, GST2.5(WT) interacts with gene 4 proteins (helicase/primase) but not GST2.5$\Delta$21C. these results proved the involvement of the carboxyl-terminal domain of gene 2.5 protein in the protein-protein interaction. We clearly conclude that carboxy-terminal domain of gene 2.5 protein is firmly involved in protein-protein interactions in T7 replication proteins.

• Food Science of Animal Resources
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• v.39 no.3
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• pp.503-509
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• 2019

The aim of this study was to compare the effects of number of washes and salt addition on the meat quality in protein recovered from Alaska Pollock compared with pork leg. Various properties of protein recovered from Alaska Pollock (C, washed twice, no salt) and pork leg (T1, washed twice, no salt; T2, two washes, salt added; T3, washed four times, no salt; and T4, washed four times, salt added) were assessed in this study. Pork leg samples exhibited better color (more whiteness, less yellowness) than Alaska Pollock samples. In pork leg samples, four washes (T3, T4) during processing yielded whiter, less yellow protein than two washes (T1, T2). Overall, the textural property measures were higher in pork leg samples (T2, T3, and T4) than in other samples. Breaking force, jelly strength, and folding resistance were significantly higher in salt-treated pork leg samples (T2, T4) than in the other samples. Our findings demonstrate that protein recovered from pork leg has better color parameters, and physical strength compared with Alaska Pollock-derived protein. A higher number of wash steps and treatment with salt during processing were furthermore found to yield better color, and physical strength in the protein samples.

• Asian-Australasian Journal of Animal Sciences
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• v.29 no.11
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• pp.1616-1624
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• 2016

The aim of present experiment was to investigate the effect of protein reduction in commercial broiler chicken rations with incorporation of de-oiled rice bran (DORB) and supplementation of limiting amino acids (valine, isoleucine, and/or tryptophan) with ration formulation either on total amino acid (TAA) or standardized ileal digestible amino acids (SIDAA). The experimental design consisted of $T_1$, TAA control; $T_2$ and $T_3$, 0.75% and 1.5% protein reduction by 3% and 6% DORB incorporation, respectively by replacing soybean meal with supplemental limiting amino acids to meet TAA requirement; $T_4$, SIDAA control, $T_5$ and $T_6$, 0.75% and 1.5% protein reduction by DORB incorporation (3% and 6%) with supplemental limiting amino acids on SIDAA basis. A total of 360 dold fast growing broiler chicks (Vencobb-400) were divided into 36 homogenous groups of ten chicks each, and six dietary treatments described were allocated randomly with six replications. During 42 days trial, the feed intake was significantly (p<0.05) reduced by TAA factor compared to SIDAA factor and protein factor significantly (p<0.05) reduced the feed intake at 1.5% reduction compared to normal protein group. This was observed only during pre-starter phase but not thereafter. The cumulative body weight gain (BWG) was significantly (p<0.05) reduced in TAA formulations with protein step-down of 1.5% ($T_3$, 1,993 g) compared to control ($T_1$, 2,067 g), while under SIDAA formulations, BWG was not affected with protein reduction of 1.5% ($T_6$, 2,076 g) compared to $T_4$ (2,129 g). The feed conversion ratio (FCR) was significantly (p<0.05) reduced in both TAA and SIDAA formulations with 1.5% protein step-down ($T_3$, 1.741; $T_6$, 1.704) compared to respective controls ($T_1$, 1.696; $T_4$, 1.663). The SIDAA formulation revealed significantly (p<0.05) higher BWG (2,095 g) and better FCR (1.684) compared to TAA formulation (2,028 g; 1.721). Intake of crude protein and all limiting amino acids (SID basis) was higher in SIDAA group than TAA group with resultant higher nitrogen retention (4.438 vs 4.027 g/bird/d). The nitrogen excretion was minimized with 1.5% protein reduction (1.608 g/bird) compared to normal protein group (1.794 g/bird). The serum uric acid concentration was significantly reduced in $T_3$ (9.45 mg/dL) as compared to $T_4$ (10.75 mg/dL). All carcass parameters were significantly (p<0.05) higher in SIDAA formulation over TAA formulation and 1.5% protein reduction significantly reduced carcass, breast and thigh yields. In conclusion, the dietary protein can be reduced by 0.75% with TAA formulation and 1.5% with SIDAA formulation through DORB incorporation and supplementation of limiting amino acids and among formulations, SIDAA formulation was better than TAA formulation.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.28 no.2
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• pp.209-218
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• 1995

Bacteriophage T7 gene 2.5 protein, a single-stranded DNA binding protein, has been implicated in T7 DNA replication, recombination, and repair. Purified gene 2.5 protein has been shown to interact with the phage encoded gene 5 protein (DNA polymerase) and gene 4 proteins (helicase and primase) and stimulates their activities. Genetic analysis of T7 phage defective in gene 2.5 shows that the gene 2.5 protein is essential for T7 DNA replication and growth. T7 phage that contain null mutants of gene 2.5 were constructed by homologous recombination. These mutant phage $(T7\Delta2.5)$ cannot grow in Escherichia coli. After infection of E. coli with $T7\Delta2.5$, host DNA synthesis is shut off, and $T7\Delta2.5$ DNA synthesis is reduced to less than $1\%$ of wild-type phage DNA synthesis (Kim and Richardson, 1993, Proc. Natl. Aca. Sci. USA, 90, 10173-10177). A truncated gene 2.5 protein $(GP2.5-\Delta21C)$ deleted the 21 carboxyl terminal amino acids was constructed by in vitro mutagenesis. $GP2.5-\Delta21C$ cannot substitute for wild-type gene 2.5 protein in vivo; the phage are not viable and exhibit less than $1\%$ of the DNA synthesis observed in wild-type phage-infected cells. $GP2.5-\Delta21C$ has been purified to apparent homogeneity from cells overexpressing its cloned gene. Purified $GP2.5-\Delta21C$ does not physically into「act with T1 gene 4 protein as measured by affinity chromatography and immunoblot analysis. The mutant protein cannot stimulate T7 gene 4 protein activity on RNA-primed DNA synthesis and primer synthesis. These results suggest that C-terminal domain of gene 2.5 protein is essential for protein-protein interactions.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.7
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• pp.1075-1079
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• 1999

Adult German cross $(German{\times}British{\times}Russian)$ angora rabbits (one year age), 32 in number were divided randomly into four groups $(T_1-T_4)$ with equal sex ratio and fed diets containing $T_1$ groundnut cake (GNC); $T_3$, soyaflakes (SF); $T_4$, sunflower cake (SFC) and $T_2$, a mixture of all the three cakes along with green forage as roughage for a period of 9 months. Nine per cent protein was added from each protein source. Fibre level was maintained by adjusting the level of rice phak in the diets. The diets were iso-nitrogenous and contained similar level of fibre. DMI through roughage was not affected due to source of protein in the diet, however, DMI through concentrate was higher $(p{\leq}0.05)$ with SFC diet, which resulted in higher total feed intake in the group $(T_4)$. Body weights increased up to second shearing, thereafter it decreased due to summer depression. Diet containing soyaflakes sustained higher wool yield whereas, it was lowest $(p{\leq}0.05)$ on SFC diet. Wool attributes (staple length, medullation, fibre diameter) were not affected due to source of protein in the diet. Digestibility of fibre and its fractions (ADF, cellulose, hemicellulose) decreased $(p{\leq}0.05)$ with incorporation of SFC in the diets. Balance of calcium was lowest whereas, that of nitrogen was highest with SFC diet $(T_4)$. Biological value of N and net protein utilization was better when different protein sources were mixed together $(T_2)$. Protein quality of soyaflakes proved better for wool production followed by groundnut cake and mixture of three protein sources. Sunflower cake alone or in combination decreased wool production which may be checked by supplementation of amino acids and energy.

• BMB Reports
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• v.37 no.2
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• pp.260-267
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• 2004

Transducin (T), the heterotrimeric guanine nucleotide binding protein in rod outer segments, serves as an intermediary between the receptor protein, rhodopsin, and the effector protein, cGMP phosphodiesterase. Labeling of T with dansyl chloride (DnsCl) inhibited its light-dependent guanine nucleotide binding activity. Conversely, DnsCl had no effect on the functionality of rhodopsin. Approximately 2-3 mol of DnsCl were incorporated per mole of T. Since fluoroaluminate was capable of activating DnsCl-modified T, this lysine-specific labeling compound did not affect the guanine nucleotide-binding pocket of T. However, the labeling of T with DnsCl hindered its binding to photoexcited rhodopsin, as shown by sedimentation experiments. Additionally, rhodopsin completely protected against the DnsCl inactivation of T. These results demonstrated the existence of functional lysines on T that are located in the proximity of the interaction site with the photoreceptor protein.

• IMMUNE NETWORK
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• v.11 no.5
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• pp.268-280
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• 2011

Background: Dengue virus, which belongs to the Flavivirus genus of the Flaviviridae family, causes fatal dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) with infection risk of 2.5 billion people worldwide. However, approved vaccines are still not available. Here, we explored the immune responses induced by alternating prime-boost vaccination using DNA vaccine, adenovirus, and vaccinia virus expressing E protein of dengue virus type 2 (DenV2). Methods: Following immunization with DNA vaccine (pDE), adenovirus (rAd-E), and/or vaccinia virus (VV-E) expressing E protein, E protein-specific IgG and its isotypes were determined by conventional ELISA. Intracellular CD154 and cytokine staining was used for enumerating CD4+ T cells specific for E protein. E protein-specific CD8+ T cell responses were evaluated by in vivo CTL killing activity and intracellular IFN-${\gamma}$ staining. Results: Among three constructs, VV-E induced the most potent IgG responses, Th1-type cytokine production by stimulated CD4+ T cells, and the CD8+ T cell response. Furthermore, when the three constructs were used for alternating prime-boost vaccination, the results revealed a different pattern of CD4+ and CD8+ T cell responses. i) Priming with VV-E induced higher E-specific IgG level but it was decreased rapidly. ii) Strong CD8+ T cell responses specific for E protein were induced when VV-E was used for the priming step, and such CD8+ T cell responses were significantly boosted with pDE. iii) Priming with rAd-E induced stronger CD4+ T cell responses which subsequently boosted with pDE to a greater extent than VV-E and rAd-E. Conclusion: These results indicate that priming with live viral vector vaccines could induce different patterns of E protein-specific CD4+ and CD8+ T cell responses which were significantly enhanced by booster vaccination with the DNA vaccine. Therefore, our observation will provide valuable information for the establishment of optimal prime-boost vaccination against DenV.

• Korean Journal of Veterinary Research
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• v.39 no.4
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• pp.786-796
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• 1999

As an attempt to develop an effective control method against theileriosis, recombinant antigen protein was produced. Thirty-two kDa membrane protein(MP) gene of T sergenti was amplified through RT-PCR from extracted total RNA of T sergenti isolated in Chonbuk, Korea. The amplified 869 bp of Korean T sergenti membrane gene was cloned and the base sequences were analyzed. The amplified gene was cloned into E coli expression vector, pQE32 plasmid vector, and the vector was introduced into E coli strain M15 to produce the recombinant membrane protein. For the induction of T sergenti membrane protein(KTs-MP), the plasmid harboring E coli strain M15 were cultured in the presence of IPTG, and the recombinant protein were purified by $Ni^+$-NTA agarose. Then, to confirm the authenticity of the produced membrane protein, molecular weight of expressed recombinant KTs-MP was analyzed by SDS-PAGE and Western blotting. The molecular weight of expressed recombinant protein was 32 kDa as expected. The recombinant KTs-MP was successfully recognized by anti-His Tag antibody, antisera of T sergenti infected cattle and monoclonal antibody of T sergenti membrane protein. Therefore, we concluded that the authentic 32 kDa membrane protein of T sergenti was produced as immunologically recognizable form.

• BMB Reports
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• v.28 no.6
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• pp.484-489
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• 1995

The product of bacteriophage T7 gene 2.5 is a single-stranded DNA binding protein and plays an important role in T7 DNA replication, recombination, and repair. Genetic analysis of T7 phage defective in gene 2.5 shows that the gene 2.5 protein is essential for T7 DNA replication and growth (Kim and Richardson, 1993). The C-terminal truncated gene 2.5 protein ($GP2.5-{\Delta}21C$) cannot substitute for wild-type gene 2.5 protein in vivo; suggesting that the C-terminal domain of gene 2.5 protein is essential for protein-protein interactions (Kim and Richardson, 1994; J. Biol. Chem. 269, 5070-5078). Truncated gene 2.5 proteins lacking 19 residues ($GP2.5-{\Delta}19N$) and 39 residues ($GP2.5-{\Delta}39N$) from the amino-terminal domain were constructed by in vitro mutagenesis. $GP2.5-{\Delta}19N$ can support the growth of T7 phage lacking gene 2.5 while $GP2.5-{\Delta}39N$ cannot substitute for wild-type gene 2.5 protein in vivo; however, its ability to bind to single-stranded DNA is not affected. These results clearly demonstrate that the 20~39 amino-terminal region of gene 2.5 protein is required for T7 growth in vivo but may not be involved in DNA binding activity.

• Korean Journal of Poultry Science
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• v.14 no.1
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• pp.25-31
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• 1987

This experiment was conducted to study the most economical and effective dietary protein level and feeding regimes in the performance of broilers. Four kinds of dietary protein level (25, 23, 20 and 18%) were prepared for this experiment. This experiment was carried on for 7 weeks and the results were shown as follows; 1. Increasing the dietary protein level resulted in a significant increase in the body weight gains (P〈0.05). 2. With a higher level of the dietary protein, the feed intake tended to be increased and the feed efficiency had same trends. 3. Increasing the dietary protein level resulted in a significant increase in the protein requirement per kg body weight gains (P〈0.05). 4. Viabilities were not affected by the different dietary protein levels. 5. The highest income had attained in the highest level of protein supplement, and the middle level of protein supplement was not higher than the lowest one. Conclusively, the highest level of protein supplement was considered to be adequate for the broiler's productiveity and income.