• BMB Reports
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• v.37 no.2
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• pp.159-166
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• 2004

Angiostatin is a potent anti-angiogenic protein. To examine the angiostatin-interacting proteins, we used the display-cloning method with a T7 phage library presenting human cDNAs. The specific T7 phage clone that bound to the immobilized angiostatin was isolated, and a novel gene encoding the displayed polypeptide on the isolated T7 phage was identified. The displayed angiostatin-binding sequence was expressed in E. coli as a soluble protein and purified to homogeneity. This novel angiostatin-binding region interacted specifically to angiostatin with a dissociation constant of $3.4{\times}10^{-7}\;M$. A sequence analysis showed that the identified sequence was a part of the large ORF of 1,998 amino acids, whose function has not yet been characterized. A Northern analysis indicated that the gene containing the angiostatin-binding sequence was expressed differentially in the developmental stages or cell types.

• Korean Journal of Microbiology
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• v.20 no.4
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• pp.210-222
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• 1982

Coliphages isolated from Han-River from September 1980 to August 1981 were classified by morphological and physiological characteristics. Effects of soil metrial on the fate of coliphage in nature were investigated. 1. The correlation coefficient between coliphage and E.coli which was host of coliphages in nature was 0.7173 (p=0.004). 2. Coliphage I isolated from Han-River of which DNA molecular weight was $27{\times}10^6$ daltons was identified as $T_1$ phage and coliphage II of which DNA molecular weight $72{\times}10^6$ daltons was classified as $T_5$ phage. 3. Soil material SW was composed of 63.65% silt and 21.92% clay. Clay was consisted of illite, kaolinite and chlorite evenly. Soil material J was composed of 68.92% silt and 11.67% clay. Clay consisted of smectite only. 4. Coliphage was absorbed to soil material J more than soil material SW, and $T_1$ coliphage was absorbed to soil material more than $T_5$ coliphage was. 5. The phage adsorption efficiency to soil material was enhanced at lower pH : the phage adsorption efficiency at pH 4 was 27 time higher than at pH 7. 6. Divalent $(Ca^{2+})\;and\;trivalention\;(Al^{3+})$ enhanced the phage adsorption efficiency to soil material from 4 to 39 and from 17 to 91 times higher than monovalent $ion(Na^+)$, respectively. 7. The concentration of organic compound was inversely related to the phage adsorption efficiency to soil. 8. Adsorption of phage onto soil material, and elution efficiency of elutants was in the order of D.D.W>tap water>river water>seawater. 9. The higher the concentration of organic compound was, the more were adsorbed phages to soil eluted. 10. Coliphages survived longer in sterile soil suspension than in nonsterile soil material suspension.

• KSBB Journal
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• v.28 no.3
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• pp.185-190
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• 2013

For the selection of peptide specifically binding to $Pb^{2+}$, the biopanning with the commercially available Ph.D.-7 phage displayed heptapeptide library was carried out against $Pb^{2+}$ immobilized on a metal-chelating IDA (iminodiacetic acid) resin. After four rounds of screening against $Pb^{2+}$-IDA including negative selections against charged bead with metal ions other than $Pb^{2+}$ and uncharged bead, several candidate lead-binding phage peptides were initially determined based on the order of frequency from the screened phage clones. Of the selected phage peptide sequences, the peptide of the highest frequency, CysSerIleArgThrLeuHisGlnCys (CSIRTLHQC) also exhibited the strongest affinity for $Pb^{2+}$ in binding assays for individual phage clones. However, there was not a significant difference in $Pb^{2+}$ affinity between selected peptides when using synthetic heptapeptides corresponding to the displayed peptide sequences of phage clones.

• BMB Reports
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• v.40 no.4
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• pp.539-546
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• 2007

The lysozymes encoded by bacteriophage T7 and K11 are both bifunctional enzymes sharing an extensive sequence homology (75%). The constructions of chimeric lysozymes were carried out by swapping the N-terminal and C-terminal domains between phage T7 and K11 lysozymes. This technique generated two chimeras, T7K11-lysozyme (N-terminal T7 domain and C-terminal K11 domain) and K11T7-lysozyme (N-terminal K11 domain and C-terminal T7 domain), which are both enzymatically active. The amidase activity of T7K11-lysozyme is comparable with the parental enzymes while K11T7-lysozyme exhibits an activity that is approximately 45% greater than the wild-type lysozymes. Moreover, these chimeric constructs have optimum pH of 7.2-7.4 similar to the parental lysozymes but exhibit greater thermal stabilities. On the other hand, the chimeras inhibit transcription comparable with the parental lysozymes depending on the source of their N-terminals. Taken together, our results indicated that domain swapping technique localizes the N-terminal region as the domain responsible for the transcription inhibition specificity of the wild type T7 and K11 lysozymes. Furthermore, we were able to develop a simple and rapid purification scheme in purifying both the wild-type and chimeric lysozymes.

• The Korean Journal of Zoology
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• v.33 no.3
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• pp.303-309
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• 1990

In order to have a handle on the availability of eukarvotic peptide hormone precursors, a cDNA encoding angler fish prepro-SRIF I was manipulated so that it can be produced in large quantity from heterologous E. coli cells. Using T7 overexpression system, fusion constructs between the T7 phage coat protein Sl0 and the prepro-SRIF were made and modified as desired. From the host E. coli strain, BL21 DE3, harboring these plasmid constructs, three different SRIF related polypeptides were expressed in large amount and characterized. The results confirm the exact construction and authenticity of the overexpressed proteins from E. coli cells. The importance of this heterologous overexpression in hard to get peptide hormone precursors as well as the suitability of the target peptide hormone SRIF for this approach are discussed.

• 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 Microbiology
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• v.27 no.4
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• pp.317-322
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• 1989

A gene can be selectively overexpressed in E. coli by utilizing the phage T7 RNA polymerase's stringent recognition and active transcription of the T7 promoter. The T7 expression system was constructed such that the T7 RNA polymerase gene is under the control of lacUV5 promoter in one plasmid, and that the target gene, the promoterless chloramphenicol acetyltransferase (CAT) gene with E. coli ribosome binding site is under the control of T7 promoter in the other plasmid. Only the E. coli cells containing both plasmids show high resistance to chloramphenicol. When the copy number of the runaway plasmid containing the polymerase gene was varied by a temperature shift, amounts of the CAT protein synthesized upon induction was correspondingly changed as shown in SDS gel electrophoresis.

• Korean Journal of Microbiology
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• v.35 no.4
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• pp.253-257
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• 1999

Effects of polyamines such as cadaverine, putrescine, spermidine and spermine on the self-splicig inhibition of the T4 phage thymidylate synthase(td) intron by spectinomycin have been investrigated. Without polyamine 7mM spectinomycin caused 40% reduction of the splicing rate. Cadaverine reduced the splicing rate over the concentrations of 0.1 to 5 mM. Putrescine at 0.5 mM increased the splicing rate by 13%. Spermidine at 0.5 mM enhanced the splicing rate by 11% while spermine at 0.01 mM enhanced the splicing rate by 16%. Of the all polyamines tested, spermine exhibited the maximum activation effect to counteract the splicing inhibition by spectinomycin. This effect appears to be due to the role of polyamine in stabilizing the conformation of td intron ribozyme essential for the catalytic function.

• Journal of Microbiology and Biotechnology
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• v.26 no.9
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• pp.1629-1635
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• 2016

A novel bacteriophage, PBES 02, infecting Cronobacter sakazakii was isolated and characterized. It has a spherical head of 90 nm in diameter and a tail of 130 nm in length, and belongs to Myoviridae as observed under a transmission electron microscope. The major virion protein appears to be 38 kilodaltons (kDa) in size. The latent period of PBES 02 is 30 min and the burst size is 250. Infectivity of the phage remained intact after exposure to temperatures ranging from 4℃ to 55℃ for 1 h. It was also stable after exposure to pHs ranging from 6 to 10 for 1 h. The phage effectively removed contaminating Cronobacter sakazakii from broth infant formula. PBES 02 has a double-stranded DNA genome of 149,732 bases. Its GC ratio is 50.7%. Sequence analysis revealed that PBES 02 has 299 open reading frames (ORFs) and 14 tRNA genes. Thirty-nine ORFs were annotated, including 24 related to replication and regulation functions, 10 related to structural proteins, and 5 related to DNA packaging. The genome of PBES 02 is closely related to that of two other C. sakazakii phages, CR3 and CR8. Comparison of DNA sequences of genes encoding the major capsid protein revealed a wide geographical distribution of related phages over Asia, Europe, and America.

• BMB Reports
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• v.31 no.5
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• pp.427-431
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• 1998

In the previous report (Choi et al., 1997), the angiogenin binding peptides identified from a phage-peptide library were analyzed by using the fusion proteins composed of the Escherichia coli maltose binding protein and its corresponding peptides. However, it was difficult to obtain a sufficient amount of the fusion proteins required for further analysis because of the low expression level. We now report a high level expression of the fusion protein and analysis of its anti-angiogenin activity. The use of strong T7 promoter and removal of signal sequence allowed about a 20-fold increase in the expression efficiency of the fusion protein. We were able to obtain about 10 mg of purified fusion protein from one liter of culture. The purified fusion protein showed angiogenin-specific affinity and inhibited the binding of biotinylated actin to human angiogenin at $IC_{50}$ of 0.6 mM. Its anti-angiogenin activity was also revealed by the chorioallantoic membrane assay.