• Korean journal of applied entomology
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• v.30 no.3
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• pp.180-186
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• 1991

This study was carried out to acquire some basic biochemical informations on the granulosis virus(GV) of Pieris rapae and Pieris brassicae. The capsule protein was composed of a single polypeptide with a molecular weight of 30,000 dalton for P. rapae GV and 31,000 dalton for P. brassicae GV. The major amino acids of capsule protein were glutamic acid, aspartic acid and lysine. When the capsule protein was partially digested with trypsin, chymotrypsin, papain or Staphylococcus aureus V8 protease, the digested products of the two viruses showed no difference in electrophoretic mobility. The patterns of the polypeptides of the two virus particle on SOS-polyacrylamide gel showed a little difference in high molecular weight region(over MW 100 kd).

• Journal of Sericultural and Entomological Science
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• no.11
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• pp.59-62
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• 1970

Many a fine structures of nuclear polyhedrosis virus in Lepidoptera had been described by electron microscope. In the larva of Antheraea pernyi Guerin, the leading virus causing infectious disease in Korea is disclosed nuclear polyhedrosis virus, which embed bundles of virus particles in the molecular lattice of polyhedra protein. The number of virus particles within a bundle. is on the average four particles, which are enclosed in a intimate membrane closely surrounded with developing membrane. The bundles of four virus particlesare at random embedded in the polyhedra protein, which is originated from the so-called virogenic stroma of chromosom in the infected nuclear.

• Journal of Life Science
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• v.19 no.5
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• pp.561-565
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• 2009

Human Tissue factor is an essential enzyme activator that forms a catalytic complex with factor VII/ VIIa, and catalyzes both the extrinsic and intrinsic blood coagulation cascades. The extracellular domain of human tissue factor is responsible for association with the biological partner. The efficient procedures for preparing biologically active human tissue factor are essential for the preclinical and clinical studies with coaguligands. An expression vector in Escherichia coli has been constructed to direct the production of extracellular human tissue factor without a fusion protein or a $His_6$ at the N-terminus. The recombinant human tissue factor was expressed in large amounts as a non-native state in E. coli. The recombinant protein was simply renatured during the DEAE-sephacel chromatographic purification procedure. Our expression and purification system does not require a protease treatment or an additional chromatographic step to remove a fusion contaminant, which provides a very useful alternative to conventional expression systems for the production of human tissue factor.

• KSBB Journal
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• v.17 no.1
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• pp.33-37
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• 2002

To improve in vitro release kinetic of G-CSF in PLGA microsphere, G-CSF was PEGylated with methoxy polyethylene glycol-aldehyde (mPEG-aldehyde, MW 5000). The majority of G-CSF was mono-PEGylated and it was characterized using SDS-PAGE, HPLC, and peptide mapping. The PLGA microencapsulation with the native, or PEGylated G-CSF was performed using W/O/w method, where the encapsulation efficiency was high. For the high loading of G-CSF to microsphere, G-CSF and PEGylated G-CSF were concentrated and then verified the protein stability using native gel and gel filtration chromatography. In comparison with native G-CSF, PEGylated G-CSF was released during the extended period and its maximum amount of released G-CSF was also increased.

• Korean Journal of Poultry Science
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• v.41 no.1
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• pp.15-20
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• 2014

Fowl adenovirus (FAV) is an important cause of several diseases, which result in considerable economic losses to the poultry farm. An outer capsid protein of FAV, fiber 2 is essential for virus growth, assembly or spread. This study was performed to produce about 22 kDa of recombinant fiber 2 protein and to immunize in laying hens to acquire the specific IgY antibody against the recombinant fiber 2. Laying hens were immunized with the recombinant fiber 2 intramuscularly in the breast muscle by injection 4 times at intervals of three weeks. At 12 weeks, serum- and egg yolk-antibody titers of hens against fiber 2 were increased up to 430,000 and 414,000, respectively. The recombinant fiber 2 could be recognized be the anti-His monoclonal antibody. Anti-fiber 2-IgY antibody could recognize the fiber 2 specifically in western blot analysis. These results suggested that the recombinant fiber 2 antigen could be used as an immunogen to elicit IgY antibody against fiber 2 and the anti-fiber 2-IgY could neutralize fowl adenovirus fiber 2 effectively.

• Microbiology and Biotechnology Letters
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• v.32 no.2
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• pp.135-141
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• 2004

Human granulocyte colony-stimulating factor (hG-CSF) is a hematopoiesis agent that principally affects the differentiation of neutrophils in the bone marrow. At present, recombinant hG-CSF is used successfully in the treatment of chemotheraphy-induced neutropenia and its indication has been expanded to bone marrow transplantation and aplastic anemia. In this study, we have constructed rhG-CSF secretion plasmid pYRC1 in which OmpA signal sequence/hG-CSF gene was expressed under the control of the T7 promoter. rhG-CSF produced in E. coli BL21 (pYRC1) grown at $37{\circ}C$ was found in aggregates. However, 15% of the periplasmic protein was soluble rhG-CSF when the E. coli BL21 (pYRC1) was cultured at $25^{\circ}C$ for 7 h in the modified MBL medium containing 10 g/$\ell$ glucose with 10 $\mu$M IPTG induction. The production of soluble rhG-CSF in E. coli BL21 (pYRC1) using fed batch culture was also studied. In the fed batch culture system, the final yield of rhG-CSF produced from E. coli BL21 (pYRC1) was increased from 4.4 mg/$\ell$to 24 mg/$\ell$by controlling the specific growth rate from $0.43 h^{-1}$ to $0.14 h^{-1}$, and optimizing the time of induction.

• KSBB Journal
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• v.23 no.3
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• pp.231-238
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• 2008

The second family member of tissue inhibitors of matrix metalloproteinases, TIMP-2, is a 21kDa protein which inhibits matrix metalloproteinases 2 (MMP-2). Expression of mammalian proteins in E. coli often forms inclusion bodies that are made up of mis-folded or insoluble protein aggregates. The requirement for the formation of 6 disulfide bonds in the process of the TIMP-2 folding is likely to be incompatible with the reducing environment of E. coli. However, this incompatibility can be often overcome by introducing a mutagenesis that could lead to enhancement of the protein solubility. In this reason, we have attempted to express the soluble TIMP-2 in E. coli by applying a modified staggered extension process (StEP), one of the in vitro PCR-based recombinant mutagenesis methods, and error-prone PCR. C-terminally located CAT fusion protein with respect to mutated TIMP-2 proteins enables us to differentiate the soluble TIMP-2 from the insoluble in E. coli by virtue of chloramphenicol resistance. According to this scheme, E. coli harboring properly-folded CAT fused to TIMP-2 protein was selected, and some of the resulting colonies exhibited an enhanced, soluble expression of TIMP-2 compared to the wild type, implying (i) the StEP technique is successfully employed to enhance the proper folding thereby increasing the solubility of TIMP-2, and (ii) the CAT dependent screening may be a simple and effective method to differentiate the soluble protein expression in E. coli.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.21 no.9
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• pp.559-568
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• 2020

Epidermal growth factor (EGF) is a hormone protein that affects cell growth and proliferation, and has various medical applications. In the present study, the gene of human EGF was codon-optimized with E. coli and the expression vector was constructed by cloning into pRSET. In order to obtain the recombinant human EGF in an active form rather than an inclusion body, chaperone co-expression was attempted along with codon optimization, for the first time. The expressed human EGF was isolated in the pure form by performing Ion Exchange Chromatography in two consecutive runs. ELISA analysis showed that the activity of purified EGF was greater than 99%, which is similar to commercially available EGF. Cell proliferation test confirmed that the recombinant human EGF has the ability to promote cell proliferation of human skin fibroblasts. The human EGF expression system of this study gives a significant amount of protein, and does not require the renaturation step and the additional chromatographic system to remove a fusion contaminant, thereby providing a very useful alternative to conventional expression systems for the preparation of recombinant human EGF.

• The Korean Journal of Malacology
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• v.22 no.1 s.35
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• pp.87-95
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• 2006

Immunohistochemical and ultrastuructural experiments were conducted to find out heavy metal accumulation in the intestine of an Antarctic gastropod Nacella concinna. According to the immune-histochemical experiment the apical cytoplasm of the intestinal epithelium showed positive reactions to anti-MT (rnotallothionein), indicating the presence of MT, a metal-binding protein involved in metal detoxifying process. In the transmission electron microscopic observations, the epithelial cells of the intestine exposed to Cd for over three hours showed irregular nuclear membranes, secretory granules, and probable metal granules. According to the SEM-EDS experiments on the intestine, concentration of Pb in the apical epithelium was in inverse proportion to that in the intestinal lumen. After exposing to Cd for over three days, S was rapidly reduced. Ca and Zn were rapidly increased after exposure to Cd. These elements are supposed to be concerned with the MT-reaction in the intestine. laken together, these data suggest that N. concinna could be used as a potential biomarker species.

• Journal of Sericultural and Entomological Science
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• v.25 no.2
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• pp.1-31
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• 1984

Flacherie, as one of the most prevalent silkworm diseases, causes severe economic damage to sericultural industry and its pathogens have been proved to be flacherie virus (FV) and densonucleosis virus (DNV). Multiplications of the viruses in the larvae of the silkworm, Bombyx mori, were studied by the sucrose density gradient centrifugation and electron microscopy. The quantitative and qualitative changes of nucleic acids and proteins were investigated from the midgut and hemolymph in the silkworm larvae infected separately with FV and DNV. The histopathological changes of epithelial cells of infected midgut also were examined by an electron microscope. 1. Purified fractions of FV or DNV in a sucrose density gradient centrifugation yielded one homogenous and sharp peak without a shoulder, suggesting no heterogenous materials in the preparation. Electron microscopy also revealed that FV and DNV were spherical particles, 27nm and 21nm in diameter, respectively. 2. Silkworm larvae showed a decrease in body weight on the 6th day and in midgut weight on the 3rd day after inoculation with FV or DNV. 3. DNA content was higher in the midgut when infected with FV or DNV, but the hemolymph of the infected larvae showed no difference during first 6 days after inoculation, after which DNA concentration declined rapidly. 4. RNA synthesis of silkworm larvae infected separately with FV and DNV was stimulated in the midgut, but RNA content was reduced in the hemolymph at the early stage of virus multiplication. At the late stage of virus multiplication, however, it was extremely reduced in both midgut and hemolymph. 5. The concentration of protein in the midgut and hemolymph of silkworm larvae infected separately with FV and DNV showed no difference from that of the healthy larvae at the early stage of virus multiplication, but it was significantly reduced at the late stage of virus multiplication. 6. There was no difference in the electrophoretic patterns of RNAs extracted from the midgut of healthy or virus-infected larvae. 7. The electrophoresis of proteins extracted from the midgut infected with FV or DNV, when carried out on the 1st and 5th day after virus inoculation, showed no difference from that of the healthy larvae. But, there was an additional band with medium motility in the proteins on the 8th day after virus inoculation, while a band with low mobility shown in the proteins of healthy larvae disappeared in the infected larvae. However, a band with high mobility in the healthy larvae was separated into two fractions in the infected larvae. 8. The electrophoretic pattern of hemolymph proteins of the silkworm larvae infected separately with FV and DNV was similar to that of the healthy larvae, but the concentration of hemolymph proteins in the infected larvae was lower than that of the healthy larvae at the late stage. 9. Two types of inclusion bodies were shown by the double staining of pyronin-methyl green in the columnar cell of the midgut on the 8th day after FV inoculation. 10. Electron microscopy of the infected midgut revealed that the 'cytoplasmic wall' of the goblet cell thickened on the 5th day after FV inoculation and several types of the cytopathogenic structures, such as virus$.$specific vesicles, virus particles, linear structures, tubular structures, and high electron-dense matrices were observed in the cytoplasm of the goblet cell. The virus particles were also observed in the microvilli and the structures similar to spherical virus particles were observed around the virus-specific vesicles, suggesting the virus assembly in the cytoplasm. 11. Fluorescence micrograph of the infected midgut stained with acridine orange showed that the nucleus, the site of DNV multiplication in the columnar cell, enlarged on the 5th day after virus inoculation. 12. Electron microscopic examination of DNV infected midgut revealed that the nucleolus of the columnar cell was broken into granules and those granules dispersed into apical region of the nucleus on the 5th day after virus inoculation. On the 8th day after inoculation, it was also observed that the nucleus of the columnar cell was full with the high electron-dense virogenic stroma which were similar to virus particles. These facts suggest that the virogenic stroma were the sites of virus assembly in the process of DNV multiplication.