• Title/Summary/Keyword: 1-2 toxin

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Disulfide Bond Bridged Divalent Antibody-Toxin, $(Fab-PE38fl)_2$ with the Toxin PE38 Fused to the Light Chain

  • Won, Jae-Seon;Choe, Mu-Hyeon
    • Journal of Microbiology and Biotechnology
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    • v.18 no.8
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    • pp.1475-1481
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    • 2008
  • B3 antibody specifically binds the $Lewis^Y$-related carbohydrate antigen of many carcinomas, and it is used as a model antibody in this study. In a previous study, the Fab fragment of the antibody was fused to a 38 kDa truncated form of Pseudomonas exotoxin A, PE38, to make Fab-PE38, where PE38 is fused to the Fd fragment of the Fab domain. This parent monomer molecule, Fab-PE38, had no cysteine in the hinge region, and it could not make a disulfide bond to form a disulfide bond bridged homodimer. In this study, we constructed three different kinds of divalent Fab-toxin fusion homodimers where the toxin is fused to the light chain of Fab, $(Fab-PE38fl)_2$. In addition to the PE38 toxin fused to the light chain, these three molecules have different hinge sequences hi, h2, and h3 making Fabh1-, Fabh2-, and Fabh3-PE38fl monomers, respectively. These hinges contain only one cysteine on different positions of the hinge sequence. The disulfide bond between the hinge region of two monomers forms homodimers $(Fabh1-PE38fl)_2$, $(Fabh2-PE38fl)_2$, and $(Fabh3-PE38fl)_2$. The refolding yields of these dimers were 5-16-fold higher than a previously constructed dimer where the PE38 was fused to the Fd fragment $(Fabh2-PE38)_2$ [8]. Our data suggest that the steric repulsion between the two PE38s in $(Fabh1-PE38)_2$ during disulfide bridge formation is relieved by fusing it at the end of the light chain. The best cytotoxicity value of these dimers showed about 2.5-fold higher on an MCF7 cell line than that of the monovalent reference molecule in ng/ml scale, which is 15-fold higher in pM scale.

Esterified-Glucomannan in Broiler Chicken Diets-Contaminated with Aflatoxin, Ochratoxin and T-2 Toxin: Evaluation of its Binding Ability (in vitro) and Efficacy as Immunomodulator

  • Raju, M.V.L.N.;Devegowda, G.
    • Asian-Australasian Journal of Animal Sciences
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    • v.15 no.7
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    • pp.1051-1056
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    • 2002
  • In vitro binding efficacy of esterified glucomannan (E-GM) (0.1%) on aflatoxin B1 (AF) (300 ppb), ochratoxin A (OA) (2 ppm) and T-2 toxin (T-2) (3 ppm), when present alone or in combination, was evaluated in toxin-contaminated feed at pH 4.5 and 6.5. Esterified glucomannan showed significantly (p<0.01) higher binding with AF (81.6%), whereas those recorded with T-2 (27.8%) and OA (25.6%) were moderate. Binding of each toxin decreased as the number of toxins in feed increased. pH of medium showed no effect on mycotoxin binding ability of E-GM. A $2{\times}2{\times}2{\times}2$ factorial experiment of 5 week duration was conducted to study the effects of two dietary levels each of AF (0 and 300 ppb), OA (0 and 2 ppm), T-2 (0 and 3 ppm ) and E-GM (0 and 0.1%) on the immune competence of a total of 960 day-old commercial broilers. Reductions in size of thymus (by AF and T-2) and bursa (by AF) and antibody titers against Newcastle disease and Infectious Bursal disease (by all the toxins) were noted. Additive and antagonistic interactions were seen among the toxins on certain parameters. Esterified glucomannan significantly (p<0.01) improved antibody titers and weights of bursa ofFabricius and thymus indicating its counteracting efficacy against immunosuppression in mycotoxicosis of multiple origin.

Isolation and Analysis of Bacillus thuringiensis serovar. darmstadiensis Insecticidal Protein Gene (Bacillus thuringiensis serovar. darmstadiensis의 곤충치사독소 유전자분리 및 구조해석)

  • 김도영;구본성;도대홍
    • The Korean Journal of Food And Nutrition
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    • v.9 no.4
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    • pp.459-465
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    • 1996
  • Bacillus thuringiensis serovar. darmstadiensis produced bipyramidal endo-toxin. The toxin protein was purified by Renografin-76 step gradient centrifugation and investigated by electron microscope. Analysis of total plasmid DNA patterns showed that four different size of plasmids existed in wild type B. thuringiensis serovar. darmstadiensis. Total plasmids DNA was isolated and transformed into pst I site of pBR322 cloning vector. Ten clones containing crystal toxin gene were forst screened colony hybridization by using PUYBT 9044 probe ontained B. thuringiensis kurskaki HD 1 toxin gene. Cloned-DNA was digested with EcoR1 and HindIII and transformed to pIBI30 sequencing vector. Finally, 2.6kb and 3.6kb size fragments contatined toxin-gene were cloned with restriction analysis.

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Evaluation of Pear Cultivar Susceptibility to AK-toxin Produced by Alternaria kikuchiana (배 검은무늬병균 (Alternaria kikuchiana)이 생성(生成)하는 AK독소(毒素)에 대한 배품종(品種)의 감수성(感受性) 진단(診斷))

  • Park, Jong Seong;Yu, Seung Hun
    • Korean Journal of Agricultural Science
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    • v.15 no.1
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    • pp.1-7
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    • 1988
  • Out of 24 isolates of Alternaria collected from pear leaves, only 7 isolates from cv. Nijiseiki leaves were specifically pathogenic to susceptible pear cultivar(Nijiseiki). Other isolates from cv. Chojuro, Oksankichi and Sinko did not show any pathogenicity to pear leaves. Pathogenic isolates of Alternaria kikuchiana produced host-specific toxin (AK-toxin) in liquid culture which caused veinal necrosis only on susceptible pear leaves, while nonpathogenic isolates did not produce this toxin. Varietal susceptibility among pear cultivars to the pathogen was investigated by evaluating HST (AK-toxin) sensitivity of pear leaves, as a substitute for spore inoculation. AK-toxin which the fungus produces was toxic to pear cultivars susceptible to the pathogen such as Isipsegi and Sinsu, but was harmless to resistant pear cultivars such as Chojuro, Oksankichi, Niitaka etc. Changes in disease susceptibility and toxin sensitivity of pear leaves with aging was investigated. Disease susceptibility and toxin sensitivity in cv. Sinsu leaves appeared to vary with leaf aging; the young leaves were visibly susceptible, but older leaves (more than 2 week old leaves) became resistant.

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Experimental Trichothecene(T-2) Toxicosis in Korean Native Goats (한국 재래산양에서의 실험적 Trichothecene(T-2) 독소중독증)

  • Kim, Jong-shu
    • Korean Journal of Veterinary Research
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    • v.28 no.1
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    • pp.49-58
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    • 1988
  • To investigate the effects of T-2 toxin on the blastogenesis of lymphocytes, pathology, hemogram and blood chemistry in the goat, the korean native goats were treated orally with T-2 toxin for 21 days with a dosage of 0.6mg per kg body weight. The results were as follows: 1. The total count of leukocytes and lymphocytes decreased significantly from 14 to 21 days after treatment. 2. Mryeloid: erythroid ratios increased significantly on days 12 after treatment. 3. Delayed-type hypersensitivity skin reactions to tuberculin were reduced predominantly. 4. T-2 toxin induced prolonged prothrombin time. 5. Mitogenic responses of lymphocytes to both lipopolysaccharide and phytohemagglutinin were significantly depressed on days 7 and 14 after treatment. 6. Treatment of T-2 toxin caused marked depletion of lymphocytes in the thymus, mesenteric lymph node, peyer's patchs and spleen.

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Experimental Trichothecene (T-2) Toxicosis in Guinea pigs (기니픽그에서의 실험적 trichothecene (T-2) 독소중독증)

  • Kim, Jong-shu;Bak, Ung-bok
    • Korean Journal of Veterinary Research
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    • v.31 no.4
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    • pp.433-440
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    • 1991
  • Guinea pigs were administrated with T-2 toxin at a rate of 1 and 0.6mg/kg body weight per day for 21 days to study the immunological and pathological effects of T-2 toxin in guinea pigs. Blood was collected before T-2 treatment and on days 7, 14 and 21 of the trial for hematological and biological examinations and for the mitogen assay using lymphocytes. Myeloid: erythroid ratios were examined from the fernur bone marrow samples taken a day before T-2 toxin treatment began, on day 12 and at death. Guinea pigs received with 1mg/kg body weight of T-2 toxin daily showed leukopenic, lymphopenic and anemic signs on day 7 and 14. The mitogenic responses to the T-cell mitogen, Concanavalin A and B-cell mitogens, lipopolysaccharide were significantly depressed on day 7. Histologically, marked cellular damages including karyorrhexis and depletion of lymphocytes were observed in the actively dividing cells of the gastrointestinal tract, lymph node, spleen and bone marrow of guinea pigs.

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Studies for Reestabilishment of Approval Toxin Amount in Paralytic Shellfish Poison-Infested Shellfish 2. Change of Toxin Composition and Specific Toxicity in Paralytic Shellfish Toxins of Blue mussel, Mytilus edulis and, Oyster, Crassostrea gigas from Woepori, $K\v{o}je$, Korea During Canning Process

  • SHIN Il-Shik;CHOI Su-Ho;LEE Tae-Sik;LEE Hi-Jung;KIM Ji-Hoe;LEE Jong-Soo;KIM Young-Man
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.29 no.6
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    • pp.900-908
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    • 1996
  • Changes of paralytic shellfish toxin components and specific toxicity in blue mussel, Mytilus edu/is and oyster, Crassostrea gigas during canning process were investigated by high performance liquid chromatography (HPLC). The $mole\%$ of the frozen shucked blue mussel were in order of $27.5\;mole\%$ of gonyautoxin 1, $23.0\;mole\%$ of gonyautoxin 8 (C1) and $23.0\;mole\%$ of epi-gonyautoxin 8 (C2), while those of the frozen shucked oyster were in order of $29\;mole\%$ of C1, $22\;mole\%$ of C2, $16.7\;mole\%$ of gonyautoxin 2. Both samples had minor amounts of saxitoxin and neosaxitoxin. On the other hand, in case of specific toxicity, the major toxins were consisted of gonyautoxin $1\~4$ in both sample. The toxicity of gonyautoxin $1\~4$ were 88 and $84\%$ in blue mussel and oyster, respectively. According to the experimental results, C1, C2 and gonyautoxin 4 were very sensitive to heat treatment, while gonyautoxin 2 and saxitoxin were pretty heat resistant than any other toxin components.

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Detection of Food Poisoning Toxin Genes Produced by Vibrio sp. (비브리오 균속이 생산하는 식중독 유발 Toxin 유전자의 검출)

  • 류병호;김민정;조경자
    • Journal of Life Science
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    • v.10 no.4
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    • pp.380-387
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    • 2000
  • Recently Vibrio sp. are the most frequently isolated microorganism, which causing food poisoning. We investigated the detection of toxin genes and effect of chitosan to toxin genes with PCR. Thirty strains of Vibrio sp. were isolated from sea water and sea products through biological and biochemical tests. Out of 30 strains, 8 were identified as V. parahemolyticus, 7 as V. mimcus, 6 as V. damsela, 5 as V. vulnificus, 4 as V. alginoyticus. In detection of ctx, tdh, and t고 as food poisoning-causing toxin genes, ctx from 7 strains, trh from 4 strains and tdh from 6 strains were detected. Among toxin genes detected strains, we selected V. vulnificus-2 V. damsela-1 and V. parahemolyticus-7. As adding chitosan solution to PCR product of 3 strains, the amplified DNA bands were not detected over 450$\mu\textrm{g}$/$m\ell$ concentrantion of chitosan. Over the result, chitosan is thought to influence the detection of toxin gene.

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Increased Refolding Yield of Disulfide Bond Bridged Fab-Toxin Homodimers by the Insertion of CH3 Domains

  • Song Jeong-Wha;Won Jae-Seon;Lee Yong-Chan;Choe Mu-Hyeon
    • Journal of Microbiology and Biotechnology
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    • v.16 no.7
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    • pp.1104-1110
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    • 2006
  • Recombinant antibody-toxin is a bifunctional protein that binds and kills a target cell expressing a specific antigen on the surface of the cell, and its structure is chimeric, in which a toxin is fused to an antigen-binding domain such as scFv or Fab. Divalent antibody-toxin molecules showed higher cytotoxicities against cancer cell lines than monovalent molecules. However, the yields of the divalent molecules were very low. In this study, we introduced the CH2, CH3, or CH2-CH3 (=Fc) domain of antibody in the middle of the Fab-toxin between the hinge region of human IgG1 and the toxin domain to increase the yield. The covalently bonded dimer could be formed by three disulfide bridges from cysteine residues in the hinge region. The molecule with the CH3 domain showed about 3-fold higher dimerization yield than previously constructed Fab-toxin molecules, while maintaining the cytotoxic activity comparable to that of scFv-toxin. However, the introduction of CH2 or Fc domain to the same position showed little effect on the dimerization yield. We also observed that the introduction of the CH3 region made it possible to form noncovalently associated dimer molecules.

Variation and Profile of Paralytic Shellfish Poisoning Toxins in Jinhae Bay, Korea

  • Mok, Jong-Soo;Song, Ki-Cheol;Lee, Ka-Jeong;Kim, Ji-Hoe
    • Fisheries and Aquatic Sciences
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    • v.16 no.3
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    • pp.137-142
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    • 2013
  • To understand critical aspects of paralytic shellfish poisoning (PSP) in a chief area of bivalve production in Korea, seasonal variation in PSP toxins in bivalves collected from Jinhae Bay, Korea in 2009 was surveyed by the pre-column high-performance liquid chromatography oxidation method. We also confirmed the profiles of major bivalves such as oysters Crassostrea gigas and mussels Mytilus galloprovincialis in Jinhae Bay. PSP toxins in the bivalves showed remarkable seasonal variation. PSP toxin levels were detected from April to May in 2009, and the highest total toxin levels at all stations were recorded in May. The major toxins in bivalves were gonyautoxin [GTX] 1&4 and C 1&2; in oysters GTX 2&3 were also detected as major components. GTX 1&4, which showed the highest PSP toxin levels at each station, accounted for the highest proportions of toxin components in mussels and oysters (64.5-71.3% and 41.4-42.4%, respectively). It was also confirmed that the highest toxicity (in ${\mu}g$ saxitoxin [STX] eq/g) was derived from GTX 1&4. The highest total toxicity (in ${\mu}g$ STX eq/g) was approximately 2-8-fold higher in mussels than in oysters collected from the same station. PSP toxin levels in bivalves differed significantly according to the sample collection station. However, the profiles of toxins in the bivalves did not show significant differences during the survey period according to sample collection station. This study shows that PSP toxin levels in some samples from Jinhae Bay were above the regulatory limit in Korea during a specific period in spring.