• Title/Summary/Keyword: permeabilization

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A Study on the Denitrification Characteristics of Permeabilized Paracoccus denitrificans (Permeabilized Paracoccus denitrificans의 탈질 특성연구)

  • 송주영;황심연;김덕술
    • KSBB Journal
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    • v.16 no.3
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    • pp.290-294
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    • 2001
  • The removal of nitrogen compounds from waste water is essential and is often accomplished by biological process. The denitrifying bacterium, Paracoccus denitrificans (KCTC 2530), was employed to study the characteristics and the denitrification differences of Permeabilized strains and untreated strains. The permeabilization rate increased with increasing toluene concentration, but some part of the toluene contributed to denaturing the datachment of proteins from the plasma membrane. Permeabilized Paracoccus denitrificans had long lag phase and high specific growth rate in cultivation, and showed excellent denitrification characteristic compared with untreated strains. But, in both cases, the denitrification ability was significantly reduced after 4 or 5 denitrifications. It seems that the strains fall into the death phase when the nutrient was exhausted. When the nutrient recovered to its initial level, the denitrification ability also recovered to the normal level. The results obtained were encouraging enough to apply to practical water treatment situation.

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Permeabilization of Ochrobactrum anthropi SY509 Cells with Organic Solvents for Whole Cell Biocatalyst

  • Park, Kyung-Oh;Song, Seung-Hoon;Yoo, Young-Je
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.9 no.3
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    • pp.147-150
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    • 2004
  • Permeabilization is known to overcome cell membrane barriers of whole cell biocatalysts. The use of organic solvents is advantageous in terms of cost, simplicity, and efficiency. In this study, Ochrobactrum anthropi SY509 was permeabilized with various organic solvents. Treatment with organic solvents resulted in lower permeability barriers due to falling out lipids of the cell membrane. Therefore, permeabilized cells showed higher enzyme activity with no cell viability. Among various organic solvents, 0.5% (v/v) chloroform was selected as the most efficient permeabilizing reagent. Changes in the cell membrane structure were observe d and the residual amounts of phospholipids of the cell membrane were measured to investigate the mechanism of the improved permeability.

Alterations in Cytoplasmic Membrane are Associated with the Bactericidal Activity of Thrombin-Induced Platelet Microbicidal Proteins in Oral Streptococci

  • Choi, Young-Eun;Cheong, Yong-Joon;Lee, Si-Young
    • International Journal of Oral Biology
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    • v.30 no.4
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    • pp.125-130
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    • 2005
  • Thrombin-induced platelet microbicidal proteins (tPMP) are antibacterial proteins released when platelets are stimulated by thrombin. It has been reported that tPMP has antibacterial activity against various bacterial species including causative agents of infective endocarditis. Most of the oral streptococci have resistance to the killing by tPMP and this fact may play an important role as a virulence factor in infective endocarditis. However, the susceptibility and resistance mechanism of oral streptococci for tPMP have not been revealed yet. In this study, the killing mechanism of tPMP for oral streptococci has been investigated. Streptococcus rattus BHT, a susceptible strain, and Streptococcus gordonii DL1, a resistant strain, have been used in this study. tPMP was isolated from platelet after stimulation with thrombin. Cell membrane depolarization was examined with 3,3'-dipropylthiodicarbocyanine iodide ($DiSC_3$), membrane potential-sensitive cyanine dye, by fluorescence spectrophotometry. The permeabilization of cell membrane by tPMP was investigated with propidium iodide (PI) by flow cytometry. tPMP susceptible S. rattus BHT showed the increase of the $DiSC_3$ fluorescence level meaning depolarization of cell membrane and increase of the uptake of PI which means permeabilization of cell membrane. However, tPMP resistant S. gordonii DLI did not show depolarization and permeabilization. These results indicate that the increasing depolarization and permeabilization of oral streptococcal cell membrane are associated with the bactericidal activity of tPMP.

Effect of Glutaraldehyde Treatment on Stability of Permeabilized Ochrobactrum anthropi SY509 in Nitrate Removal

  • Park, Young-Tae;Park, Jae-Yeon;Park, Kyung-Moon;Choi, Suk-Soon;Yoo, Young-Je
    • Journal of Microbiology and Biotechnology
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    • v.18 no.11
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    • pp.1803-1808
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    • 2008
  • For practical application, the stability of permeabilized Ochrobactrum anthropi SY509 needs to be increased, as its half-life of enzymatic denitrification is only 90 days. As the cells become viable after permeabilization treatment, this can cause decreased activity in a long-term operation and induce breakage of the immobilization matrix. However, the organic solvent concentration causing zero cell viability was 50%, which is too high for industrial application. Thus, whole-cell immobilization using glutaraldehyde was performed, and 0.1% (v/v) glutaraldehyde was determined as the optimum concentration to maintain activity and increase the half-life. It was also found that 0.1% (v/v) glutaraldehyde reacted with 41.9% of the total amine residues on the surface of the cells during the treatment. As a result, the half-life of the permeabilized cells was increased from 90 to 210 days by glutaraldehyde treatment after permeabilization, and no cell viability was detected.

Continuous Production of Gluconic Acid and Sorbitol from Glucose and Fructose using Perrneabilized cells of Zymomonas mobilis (투과화된 Zymomonas mobilis 균체를 이용한 Glucose와 Fructose로 부터 Gluconic Acid와 Sorbitol의 생산)

  • 김원준;박제균;김학성
    • Microbiology and Biotechnology Letters
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    • v.19 no.1
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    • pp.70-75
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    • 1991
  • Continuous and simultaneous production of gluconic acid and sorbitol from glucose and fructose was carried out by using glucose-fructose oxidoreductase and glucanolactonase of Zymomonas mobilis. In order to utilize the enzymes without purification, Zymomonas mobilis was permeabilized with toluene. Optimum conditions for permeabilization and reaction kinetics of permeabilized Zymomonas mobilis were studied. In batch operation with the permeabilized cells immobilized in alginate beads, about 90% conversion was obtained within 35 h reaction. Continuous production of gluconic acid and sorbitol using the immobilized permeabilized cells was carried out. Optimum conditions for continuous operation with the imn~obilized cells were; pH 6.2 and temperature $40^{\circ}C$. Maximum productivities for gluconic acid and sorbitol were about 14.5 g/l/h and 14.8 g/l/h respectively at the dilution rate of 0.075 $h^{-1}$ when 300 g/l each of substrates was fed.

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Screening and Purification of an Antimicrobial Peptide from the Gill of the Manila Clam Ruditapes philippinarum (바지락(Ruditapes philippinarum) 아가미로부터 항균 펩타이드의 탐색 및 정제)

  • Seo, Jung-Kil
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.49 no.2
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    • pp.137-145
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    • 2016
  • This study screened the biological activity of an acidified gill extract of the Manila clam Ruditapes philippinarum including antimicrobial, hemolytic, membrane permeabilization, and DNA-binding activity, and purified the antimicrobial material. The acidified gill extract showed potent antimicrobial activity against Bacillus subtilis and Escherichia coli without significant hemolytic activity, but showed no membrane permeabilization or DNA-binding ability. An antimicrobial material was purified from the acidified gill extract using C18 reversed-phase and cation-exchange high-performance liquid chromatography (HPLC). Treatment of the purified material with trypsin completely abolished all of the antibacterial activity against Bacillus subtilis, suggesting that the purified material is a proteinaceous antibiotic. The molecular weight of the purified material was 2571.9 Da, but no primary structural information was obtained due to N-terminal blocking. A future study should confirm the primary structure. Our results suggest that the Manila clam gill contains proteinaceous antibiotics that have a role in first-line defense. This information could be used to better understand the Manila clam innate immune system.

Biphasic Activity of Chloroquine in Human Colorectal Cancer Cells

  • Park, Deokbae;Lee, Youngki
    • Development and Reproduction
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    • v.18 no.4
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    • pp.225-231
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    • 2014
  • Autophagy is a homeostatic degradation process that is involved in tumor development and normal development. Autophagy is induced in cancer cells in response to chemotherapeutic agents, and inhibition of autophagy results in enhanced cancer cell death or survival. Chloroquine (CQ), an anti-malarial drug, is a lysosomotropic agent and is currently used as a potential anticancer agent as well as an autophagy inhibitor. Here, we evaluate the characteristics of these dual activities of CQ using human colorectal cancer cell line HCT15. The results show that CQ inhibited cell viability in dose- and time-dependent manner in the range between 20 to 80 uM, while CQ did not show any antiproliferative activity at 5 and 10 uM. Cotreatment of CQ with antitumor agent NVP-BEZ235, a dual inhibitor of PI3K/mTOR, rescued the cell viability at low concentrations meaning that CQ acted as an autophagy inhibitor, but CQ induced the lethal effect at high concentrations. Acridine orange staining revealed that CQ at high doses induced lysosomal membrane permeabilization (LMP). High doses of CQ produced cellular reactive oxygen species (ROS) and cotreatment of antioxidants, such as NAC and trolox, with high doses of CQ rescued the cell viability. These results suggest that CQ may exert its dual activities, as autophagy inhibitor or LMP inducer, in concentration-dependent manner.

Enhanced Activity of Phenylalanine Ammonia Lyase in Permeabilised Recombinant E. coli by Response Surface Method

  • Cui, Jian-dong;Li, Yan;Jia, Shi-Ru
    • Food Science and Biotechnology
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    • v.18 no.2
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    • pp.494-499
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    • 2009
  • To improve phenylalanine ammonia lyase (E.C.4.3.1.5-PAL) activity in recombinant Escherichia coli, Some approaches for improving phenylalanine ammonia lyase (PAL) activity in recombinant E. coli were developed following preliminary studies by means of response surface method. The results shown that permeabilization with combination of Triton X-100, cetyl trimethyl ammonium bromide (CTAB), and acetone enriched cellular recombinant PAL activity significantly, which improved over 10-fold as compared with the control (untreat cell), as high as 181.37 U/g. The optimum values for the tested variables were Triton X-100 0.108 g/L, CTAB 0.15 g/L, and acetone 45.2%(v/v). Furthermore, a second-order model equation was suggested and then validated experimentally. It was indicated that addition of surfactants and organic solvents made the cells more permeable and therefore allowed easier access of the substrate to the enzyme and excretion of the product, which increased the rate of transport of L-phenylalanine and trans-cinnamic acids. These improved methods of PAL activity enrichment could serve as a rich enzyme source, especially in the biosynthesis of L-phenylalanine.

Aucklandia lappa Causes Membrane Permeation of Candida albicans

  • Lee, Heung-Shick;Kim, Younhee
    • Journal of Microbiology and Biotechnology
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    • v.30 no.12
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    • pp.1827-1834
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    • 2020
  • Candida albicans is a major fungal pathogen in humans. In our previous study, we reported that an ethanol extract from Aucklandia lappa weakens C. albicans cell wall by inhibiting synthesis or assembly of both (1,3)-β-D-glucan polymers and chitin. In the current study, we found that the extract is involved in permeabilization of C. albicans cell membranes. While uptake of ethidium bromide (EtBr) was 3.0% in control cells, it increased to 7.4% for 30 min in the presence of the A. lappa ethanol extract at its minimal inhibitory concentration (MIC), 0.78 mg/ml, compared to uptake by heat-killed cells. Besides, leakage of DNA and proteins was observed in A. lappa-treated C. albicans cells. The increased uptake of EtBr and leakage of cellular materials suggest that A. lappa ethanol extract induced functional changes in C. albicans cell membranes. Incorporation of diphenylhexatriene (DPH) into membranes in the A. lappa-treated C. albicans cells at its MIC decreased to 84.8%, after 60 min of incubation, compared with that of the controls, indicate that there was a change in membrane dynamics. Moreover, the anticandidal effect of the A. lappa ethanol extract was enhanced at a growth temperature of 40℃ compared to that at 35℃. The above data suggest that the antifungal activity of the A. lappa ethanol extract against C. albicans is associated with synergistic action of membrane permeabilization due to changes in membrane dynamics and cell wall damage caused by reduced formation of (1,3)-β-D-glucan and chitin.