• Title/Summary/Keyword: proteinase K

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Influence of Growth Conditions for the Production of Bacteriocin, Glycinecin, Produced by Xanthmonas campestris pv. glycines 8ra (콩 불마름병균의 생장 조건이 박테리오신인 glycinecin의 생성에 미치는 영향)

  • Woo Jung;Sunggi Heu;Cho, Yong-Sup
    • Korean Journal Plant Pathology
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    • v.14 no.5
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    • pp.376-381
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    • 1998
  • Xanthomonas campestris pv. glycines 8ra causes bacterial pustule disease on susceptible soybean leaves and produces a bacteriocin, named glycinecin, against related bacteria such as Xanthomonas campestris pv. vesicatoria. The antimicrobial activity of the glycinecin was effective to most tested Xanthomonas species. X. c. pv. glycines 8ra was able to produce the glycinecin in liquid media as well as solid media. Maximal productivity of glycinecin was obtained at 3$0^{\circ}C$ in the early stationary phase of growth of the X. c. pv. glycines 8ra. The production of glycinecin was not dependent on the initial inoculum level but on cell density. Glycinecin was very sensitive to proteolytic enzymes such as trypsin and proteinase K but resistant to DNase and RNase. The culture supernatant of X. c. pv. glycines 8ra retained some of its antimicrobial activity after 15 min at 6$0^{\circ}C$. It is stable at wide range of pH. The glycinecin showed the bactericidal activity after the adsorption of the glycinecin to the sensitive bacterial cell.

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Changes in Profiles of Major Proteins in Encysting Acanthamoeba castellanii

  • Park, Joon-Tae;Jeong, Young-Eui;Ahn, Tae-In
    • Animal cells and systems
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    • v.6 no.4
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    • pp.341-347
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    • 2002
  • The life cycle of Acanthamoeba is comprised of two distinct stages, tropho-zoite and cyst. During periods of stress, trophozoites undergo cellular differentiation into cyst. In order to understand the cellular differentiation, ore followed changes in profiles of major proteins by 2D-PAGE and ubiqui-tinated proteins by immunoblotting with anti-ubiquitin (Ub) monoclonal antibody (mAb) as a probe. We observed 51 proteins present in trophozoite were lost with the encystment. We found that 43 proteins within 24 h, and 8 proteins in 96 h of encystment. Among them, 17 proteins were staine with anti-Ub mAb. In cysts, 16 proteins including 2 anti-Ub mAb-reactive proteins were newly synthesized. Four proteins were newly detected in 24 h-cyst and disappeared in 96 h-cyst, one protein was synthesized in 24-96 h-cyst and disappeared in 168 h-cyst, and 11 proteins appeared upon en-cystment and were present in all cyst stages. We identified a cyst specific 33 kDa protein as subtilisin-like serine proteinase by N-terminal sequencing. Identification of these proteins lost and newly synthesized with encystment would improve our understanding of cysting protozoan parasites.

Biodegradation of Organophosphate Pesticide Using Recombinant Cyanobacteria with Surface- and Intracellular-Expressed Organophosphorus Hydrolase

  • Chungjatupornchai, Wipa;Fa-Aroonsawat, Sirirat
    • Journal of Microbiology and Biotechnology
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    • v.18 no.5
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    • pp.946-951
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    • 2008
  • The opd gene, encoding organophosphorus hydrolase (OPH) from Flavobacterium sp. capable of degrading a wide range of organophosphate pesticides, was surface- and intracellular-expressed in Synechococcus PCC7942, a prime example of photoautotrophic cyanobacteria. OPH was displayed on the cyanobacterial cell surface using the truncated ice nucleation protein as an anchoring motif. A minor fraction of OPH was displayed onto the outermost surface of cyanobacterial cells, as verified by immunostaining visualized under confocal laser scanning microscopy and OPH activity analysis; however, a substantial fraction of OPH was buried in the cell wall, as demonstrated by proteinase K and lysozyme treatments. The cyanobacterial outer membrane acts as a substrate (paraoxon) diffusion barrier affecting whole-cell biodegradation efficiency. After freeze-thaw treatment, permeabilized whole cells with intracellular-expressed OPH exhibited 14-fold higher bioconversion efficiency ($V_{max}/K_m$) than that of cells with surface-expressed OPH. As cyanobacteria have simple growth requirements and are inexpensive to maintain, expression of OPH in cyanobacteria may lead to the development of a low-cost and low-maintenance biocatalyst that is useful for detoxification of organophosphate pesticides.

New anti-wrinkle cosmetics

  • Lee, Kang-Tae;Lee, Sun-Young;Jeong, Ji-Hean;Jo, Byoung-Kee
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.28 no.1
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    • pp.71-79
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    • 2002
  • In the aged skin especially in the face and eyelid, deep and slight wrinkles are one of the remarkable phenomena of aging and the cause of wrinkle is various. Among the cause of wrinkles an oxidative stress plays an important roles in wrinkle formation process. It caused the lipid peroxidation of cell membrane, the increase of the MMPs(MatrixMetalloProteinase) gene expression and cellular DNA damage. These ROS induced materials may cause the degradation of collagen matrix system in the dermis and cause the formation of skin wrinkle. So, it is very important for protecting skin wrinkle formation to regulate ROS activity. In this study, we developed one active ingredient having multi functional activities such as activation of collagen synthesis, inhibition of MMPs activity, lipid peroxidation and free radical scavenging activity and inhibition of free radical induced DNA damage in vitro. Pericarpium castaneae extracts showed collagen synthesis increase in Normal Human Fibroblast and the inhibition of elastase activity (IC$\_$50/ of Elastase: 43.9$\mu\textrm{g}$/㎖). It showed also anti-oxidative activity (IC$\_$50/ : 48$\mu\textrm{g}$/㎖) and free radical scavenging activity(IC$\_$50/: 7.6$\mu\textrm{g}$/㎖). Conclusively, Pericarpium castaneae extracts may be used as an ingredient for new anti-wrinkle cosmetics.

Identification of Antifungal Substances of Lactobacillus sakei subsp. ALI033 and Antifungal Activity against Penicillium brevicompactum Strain FI02

  • Huh, Chang Ki;Hwang, Tae Yean
    • Preventive Nutrition and Food Science
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    • v.21 no.1
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    • pp.52-56
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    • 2016
  • This study was performed to investigate the antifungal substances and the antifungal activity against fungi of lactic acid bacteria (LAB) isolated from kimchi. LAB from kimchi in Imsil showed antifungal activity against Penicillium brevicompactum strain FI02. LAB LI031 was identified as Lactobacillus sakei subsp. Antifungal substances contained in L. sakei subsp. ALI033 culture media were unstable at high pH levels. Both, the control and proteinase K and protease treated samples showed clear zones, suggesting that the antifungal substances produced by ALI033 were non-protein substances unaffected by protesases. Both, the control and catalase showed clear zones, suggesting that the antifungal metabolite was not $H_2O_2$. The molecular weights of the antifungal substances were ${\leq}3,000Da$. The organic acid content of crude antifungal substances produced by L. sakei subsp. ALI033 showed high concentrations of lactic acid (502.47 mg/100 g). Therefore, these results suggest that antifungal substance produced by L. sakei subsp. ALI033 is most likely due to its ability in producing organic acid.

Potential Role of Bacterial Infection in Autoimmune Diseases: A New Aspect of Molecular Mimicry

  • Alam, Jehan;Kim, Yong Chul;Choi, Youngnim
    • IMMUNE NETWORK
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    • v.14 no.1
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    • pp.7-13
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    • 2014
  • Molecular mimicry is an attractive mechanism for triggering autoimmunity. In this review, we explore the potential role of evolutionary conserved bacterial proteins in the production of autoantibodies with focus on granulomatosis with polyangiitis (GPA) and rheumatoid arthritis (RA). Seven autoantigens characterized in GPA and RA were BLASTed against a bacterial protein database. Of the seven autoantigens, proteinase 3, type II collagen, binding immunoglobulin protein, glucose-6-phosphate isomerase, ${\alpha}$-enolase, and heterogeneous nuclear ribonuclear protein have well-conserved bacterial orthologs. Importantly, those bacterial orthologs are also found in human-associated bacteria. The wide distribution of the highly conserved stress proteins or enzymes among the members of the normal flora and common infectious microorganisms raises a new question on how cross-reactive autoantibodies are not produced during the immune response to these bacteria in most healthy people. Understanding the mechanisms that deselect auto-reactive B cell clones during the germinal center reaction to homologous foreign antigens may provide a novel strategy to treat autoimmune diseases.

Purification and Characterization of an Antibacterial Substance from Aerococcus urinaeequi Strain HS36

  • Sung, Ho Sun;Jo, Youl-Lae
    • Journal of Microbiology and Biotechnology
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    • v.30 no.1
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    • pp.93-100
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    • 2020
  • A bacterial strain inhibiting the growth of Vibrio anguillarum, the causative agent of vibriosis, was isolated from fish intestines. The isolated strain HS36 was identified as Aerococcus urinaeequi based on the characteristics of the genus according to Bergey's Manual of Systematic Bacteriology and by 16S rRNA sequencing. The growth rate and antibacterial activity of strain HS36 in shaking culture were higher than those in static culture, while the optimal pH and temperature for antibacterial activity were 7.0 and 30℃, respectively. The active antibacterial substance was purified from a culture broth of A. urinaeequi HS36 by Sephadex G-75 gel chromatography, Sephadex G-25 gel chromatography, and reverse-phase high-performance liquid chromatography. Its molecular weight, as estimated by Tricine SDS-polyacrylamide gel electrophoresis, was approximately 1,000 Da. The antibacterial substance produced by strain HS36 was stable after incubation for 1 h at 100℃. Although its antibacterial activity was optimal at pH 6-8, activity was retained at a pH range from 2 to 11. The purified antibacterial substance was inactivated by proteinase K, papain, and β-amylase treatment. The newly purified antibacterial substance, classified as a class II bacteriocin, inhibited the growth of Klebsiella pneumoniae, Salmonella enterica, and Vibrio alginolyticus.

Involvement of Cathepsin D in Apoptosis of Mammary Epithelial Cells

  • Seol, M.B.;Bong, J.J.;Baik, M.
    • Asian-Australasian Journal of Animal Sciences
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    • v.19 no.8
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    • pp.1100-1105
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    • 2006
  • During involution of the mammary gland after the lactation period, the gland undergoes an extensive epithelial cell death. In our previous study, overexpression of an extracellular proteinase inhibitor (Expi) gene accelerated apoptosis of mammary epithelial cells. Here we found that expression of the cathepsin D gene was induced in the Expi-overexpressed apoptotic cells. To understand the role of cathepsin D in apoptosis, we transfected cathepsin D gene into mammary epithelial HC11 cells and established the stable cell lines overexpressing the cathepsin D gene. We found that overexpression of the cathepsin D gene partially induced apoptosis of mammary epithelial cells. Expression patterns of the cathepsin D gene were examined in mouse mammary gland at various reproductive stages. Expression of the cathepsin D gene was increased during involution stages compared to lactation stages, and highest expression levels were shown at involution on day 4. We also examined expression of the cathepsin D gene in various mouse tissues. Mammary gland at involution on day 2 showed highest levels of cathepsin D mRNA of the mouse tissues that we examined. Liver tissues showed high levels of cathepsin D expression. These results demonstrate that cathepsin D may contribute to the apoptotic process of mammary epithelial cells.

Increased Association of ${\alpha}$-synuclein to Perturbed Cellular Membranes

  • Kim, Yoon-Suk;Lee, Seung-Jae
    • Biomedical Science Letters
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    • v.17 no.2
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    • pp.167-171
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    • 2011
  • [ ${\alpha}$ ]synuclein (${\alpha}$-syn) is implicated in the pathogenesis of Parkinson's disease (PD) and other related diseases. We have previously reported that ${\alpha}$-syn binds to the cell membranes in a transient and reversible manner. However, little is known about the physiologic function and/or consequence of this association. Here, we examined whether chemically induced perturbations to the cellular membranes enhance the binding of ${\alpha}$-syn, based on hypothesis that ${\alpha}$-syn may play a role in maintenance of membrane integrity or repair. We induced membrane perturbations or alterations in ${\alpha}$-syn-overexpressing human neuroblastoma cells (SH-SY5Y) by treating the cells with hydrogen peroxide ($H_2O_2$) or oleic acid. In addition, membranes fractionated from these cells were perturbed by treating them with proteinase K or chloroform. Dynamic interaction of ${\alpha}$-syn to the membranes was analyzed by the chemical cross-linking assay that we developed in the previous study. We found that membrane interaction of ${\alpha}$-syn was increased upon treatment with membrane-perturbing reagents in a dose and time dependent manner. These results suggest that perturbations in the cellular membranes cause increased binding of ${\alpha}$-syn, and this may have significant implication in the physiological function of ${\alpha}$-syn in cells.

Characterization of Bacteriocin Produced by Enterococcus faecium MJ-14 Isolated from Meju

  • Lim, Sung-Mee;Park, Mi-Yeon;Chang, Dong-Suck
    • Food Science and Biotechnology
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    • v.14 no.1
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    • pp.49-57
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    • 2005
  • Enterococcus faecium MJ-14, having strong antilisterial activity, was isolated from Korean fermented food, Meju. MJ-14 showed the same phenotypic characteristics, but different sugar utilization, as reference strain, E. faecium KCCM12118. It could utilize D-xylose, amygdaline, and gluconate, whereas E. faecium KCCM12118 could not. Optimal condition for bacteriocin production by E. faecium MJ-14 was at $37^{\circ}C$ and pH 7.0. Bacteriocin activity appeared in mid exponential phase and increased rapidly up to stationary phase. Activity was significantly promoted in MRS broth containing 3.0% glucose, 1.5% lactose, 2.0% peptone, or 1.5% tryptone. Bacteriocins effectively inhibited Enterococcus faecalis and Listeria spp. of Gram-positive bacteria, and Helicobacter pylori of Gram-negative bacteria, but did not inhibit yeasts and molds. They were stable against heat (for 30 min at $100^{\circ}C$), pH (3.0-9.0), long-term storage (for 60 days at 4 or $-20^{\circ}C$), and enzymatic digestion by catalase, proteinase K, papain, lysozyme, trypsin, chymotrypsin, and lipase, etc. Bacteriocin activity was completely inhibited by protease and pepsin, and 50% by ${\alpha}$-amylase. Studies on PCR detection of enterocin structural genes revealed bacteriocins are identical to enterocins A and B.