• Title/Summary/Keyword: affinity binding

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Characterization of the Four GH12 Endoxylanases from the Plant Pathogen Fusarium graminearum

  • Habrylo, Olivier;Song, Xinghan;Forster, Anne;Jeltsch, Jean-Marc;Phalip, Vincent
    • Journal of Microbiology and Biotechnology
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    • v.22 no.8
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    • pp.1118-1126
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    • 2012
  • Four putative GH12 genes were found in the Fusarium graminearum genome. The corresponding proteins were expressed in Escherichia coli, purified, and evaluated. FGSG_05851 and FGSG_11037 displayed high activities towards xyloglucan ($V_{max}$ of 4 and $11{\mu}mol/min$, respectively), whereas FGSG_07892 and FGSG_16349 were much less active with this substrate (0.081 and $0.004{\mu}mol/min$, respectively). However, all four of these enzymes had a similar binding affinity for xyloglucan. Xyloglucan was the substrate preferred by FGSG_05851, in contrast to the three other enzymes, which preferred ${\beta}$-glucan or lichenan. Therefore, FGSG_05851 is a xyloglucan-specific glucanase (E.C. 3.2.1.151) rather than an endoglucanase (E.C. 3.2.1.4) with broad substrate specificity. FGSG_11037 displayed a peculiar behavior in that the xyloglucan binding was highly cooperative, with a Hill coefficient of 2.5. Finally, FGSG_05851 essentially degraded xyloglucan into hepta-, octa-, and nonasaccharides, whereas the three other enzymes yielded hepta- and octa-saccharides as well as larger molecules.

Characterization of a Substance from Photobacterium damsela subsp. piscicida that Non-specifically Binds to Streptavidin

  • Jung Tae Sung;Kim D. Thompson;Adams Aelexandra;Oh Myung Joo
    • Fisheries and Aquatic Sciences
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    • v.3 no.1
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    • pp.52-63
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    • 2000
  • Non-specific reaction has been a problem in doing, especially, research and diagnosis for infectious agents. Avidin-biotin-peroxidase complex (ABC) techniques has widely been used to amplify a reaction. Photobacterium damse1a subsp. piscicdia (formerly Pasteurella piscicida) exhibited a capacity to bind with streptavidin non-specifically. The band, estimated 26 K Da in Western blotted paper, was blocked with biotin but incompletely. In an attempt to explore an involvement of the non-specific substance in attaching piscine cells, cell attachment test performed using anti- Ph. d. subsp piscicida sera raised mouse and rabbit exhibited slightly blocking effects for Mediterranean (1736) and significantly for Japanese (Sp 92144) isolate. Biotin decreased the attachment ability significantly for Sp92144 but it was not effective to 1736. Both isolates showed greatly enhanced attachment ability with poly-L-lysin. The non-specific binding substance was contained in bacterial extracellular products (ECPs). The substance was able to purified with 2-imminobiotin affinity column, the purified substance appeared to have 4 bands in silver staining, and had a carbohydrate branch. This purified substance showed cytotoxic effects selectively between 5 piscine cell lines. Moreover, it stimulated rainbow trout macrophage in terms of reduction of cytochrome cas well as yeast phagocytosis, significantly.

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Antimicrobial Flavonoid, 3,6-Dihydroxyflavone, Have Dual Inhibitory Activity against KAS III and KAS I

  • Lee, Jee-Young;Lee, Eun-Jung;Jeong, Ki-Woong;Kim, Yang-Mee
    • Bulletin of the Korean Chemical Society
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    • v.32 no.9
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    • pp.3219-3222
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    • 2011
  • Three types of ${\beta}$-ketoacyl acyl carrier protein synthase (KAS) are important for overcoming the bacterial resistance problem. Recently, we reported the discovery of a antimicrobial flavonoid, YKAF01 (3,6-dihydroxyflavone), which exhibits antibacterial activity against Gram-positive bacteria through inhibition of ${\beta}$-ketoacyl acyl carrier protein synthase III (KAS III). In this report, we suggested that YKAF01 can be an inhibitor ${\beta}$-ketoacyl acyl carrier protein synthase I (KAS I) with dual inhibitory activity for KAS I as well as KAS III. KAS I is related to the elongation of unsaturated fatty acids in bacterial fatty acid synthesis and can be a good therapeutic target of designing novel antibiotics. We performed docking study of Escherichia coli KAS I (ecKAS I) and YKAF01, and determined their binding model. YKAF01 binds to KAS I with high binding affinity ($2.12{\times}10^6$) and exhibited an antimicrobial activity against the multidrug-resistant E. coli with minimal inhibitory concentration (MIC) value of 512 ${\mu}g$/mL. Further optimization of this compound will be carried out to improve its antimicrobial activity and membrane permeability against bacterial cell membrane.

(A) Study on Natural Dyeing of polyester(I) - Ginseng Microcapsules - (폴리에스테르의 천연염색 처리 방법에 관한 연구(I) - 인삼 마이크로캡슐을 중심으로 -)

  • Min, Kyung-Hae
    • Fashion & Textile Research Journal
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    • v.10 no.4
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    • pp.560-565
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    • 2008
  • Plantae of the natural material has been broadly used on cloth dyeing by it's specific properties such as eco-friendly and innoxious. However dyeing with natural material on synthetic fiber is nearly impossible due to poor affinity between natural material and synthetic fiber. The method which is binding with micro-capsulized natural material to cloth, used in this study, has low change on quality by external influence. Also this method has high ability in spray effect by broken capsule which comes to pressure and friction when the treated cloth was dressed. And this method is applicable widely from natural fiber to synthetic fiber. The purpose of this study is to develop the multi-functional synthetic material with micro-capsulized Ginseng on PET. Moreover, it was driven by comparison of colormetric properties and fastness between regular dip-dyeing method and binding with micro-capsulized material method. Dyeability showed a little bit low exhaustion but the PET treated by micro-capsule was more or less better than the dip dyed PET. Through the SEM(Scanning Electron Microscope) of PET treated by micro-capsule, it has good residence of capsules even after 5 or 10 times washing. Wash and light fastness were revealed some different grades by each condition but showed high level, in most and the micro-capsulized PET was more improver than regular dip dyed PET.

β-Secretase (BACE1) Purification by Refolding Method and Complex with Hispidin

  • Lim, Ji-Hong;Lee, Bo Ram;Park, Hee Won;Hong, Bum Soo;Lim, Beong Ou;Kim, Young Jun
    • Journal of the Korean Chemical Society
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    • v.58 no.6
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    • pp.553-559
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    • 2014
  • Alzheimer's disease (AD) is a devastating neurodegenerative disease that represents the most common form of dementia among the elderly population. The deposition of aggregated ${\beta}$-amyloid ($A{\beta}$) senile plaques in the human brain is a classic observation in the neuropathology of AD, yet an understanding of the mechanism of their formation remains elusive. $A{\beta}$ is formed through endoproteolysis of the amyloid precursor protein (APP) by ${\beta}$-secretase (BACE1, ${\beta}$-site APP-cleaving enzyme) and ${\gamma}$-secretase. In this study, BACE1 protein was successfully over-expressed, purified, and refolded and utilized in a binding study with hispidin. We developed a simpler refolding method using a urea gradient and size-exclusion gel filtration to purify an active BACE1 protein variant, in larger quantities than that reported previously, and measured the binding affinity of hispidin to the BACE1 protein variant through isothermal titration calorimetry.

Prediction of HLA-A*0201-Restricted Antigenic Epitopes Targeting Multiple Myeloma (다발성 골수종 적용을 위한 HLA-A*0201 제한 항원성 펩타이드 예측)

  • Kang, Yoon Joong
    • Journal of Convergence for Information Technology
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    • v.10 no.6
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    • pp.209-216
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    • 2020
  • Protein antigens and their epitopes are targets for epitope based vaccines. There are many prediction servers which can be used for identification of binding peptides to MHC molecules. However, choosing of appropriate prediction servers is difficult. This study compared data obtained from prediction servers and evaluate them in scope of binding affinity to MHC-I molecules. Here we predicted HLA-A2-restricted cytotoxic T lymphocyte epitopes from survivin as a potential target for multiple myeloma. We suggest a procedure for prediction of antigenic peptides which could bind to MHC-I molecule. The results of this study will assist researchers in selection and prediction of noble antigenic peptides.

Ig G fusion 단백질을 사용한 리간드-수용체의 상호작용

  • 천혜경
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1994.11a
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    • pp.143-145
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    • 1994
  • Chimeric fusion proteins involving IgG have proven valuable in studying protein-protein interactions and may possess therapeutic applications as well. For example, three receptor subtypes for the natriuretic peptides, when fused to the Fc portion of human IgG ${\gamma}$ chain, were quantitatively and qualitatively indistinguishable from the native receptor, thus allowing detailed structure-function studies of the receptor. In an attempt to block human immunodeficiency virus infectivity with soluble derivatives of CD4, a CD4/IgG Fc chimeric molecule was shown to increase the plasma half life of soluble CD4 and possessed the added advantage of IgG Fc-mediated placental transfer. In the case of the KGFR, this approach provided a framework for dissection of its ligand binding domains and made it possible to demonstrate that high affinity binding sites for two ligands, aFGF and KGF, reside within different receptor Ig-like domains. Chimeric molecules fused to immunoglobulins would have the advantages of secretion from transfected cells as well as detection and purification from medium utilizing Staphylococcus aureus Protein A. In addition, where highly related receptors make their discrimination very hard due to the difficulties in generating specific immunochemical probes, IgG fusion protein with tailor-made specificities confers particular advantages to elucidate patterns of receptor distribution and expression. The approach described here may have general applications in defining ligand-receptor interactions as well as searching for specific agonists and antagonists of receptor function.

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Field Study on the Mycotoxin Binding Effects of Clay in Oreochromis niloticus Feeds and Their Impacts on the Performance as Well as the Health Status throughout the Culture Season

  • Abdelaziz, Mohamed;Anwer, Wael;Abdelrazek, Abeer Hamada
    • Interdisciplinary Bio Central
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    • v.2 no.4
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    • pp.10.1-10.5
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    • 2010
  • Total aflatoxin and ochratoxin were detected in 3 naturally contaminated fish feed samples using immune-affinity method. The results revealed that the average levels of aflatoxins in the 3 examined samples were (15, 22 and $12\;{\mu}g/kg$) respectively while the average levels of ochratoxins were (15, 6 and $6\;{\mu}g/kg$). The results of determination of the effects of clay as a mycotoxin binder on the health status and performance of Oreochromis niloticus in comparing with a control group revealed that the survival rate in control group was 81% after the end of the culture season. The results also revealed that the survival rate in group 2 which received clay treated feed was 86%. The results of regular parasitological examination revealed the identification of trichodina as external protozoa in Oreochromis niloticus from both ponds but without manifestation of disease signs. The results of bacteriological examination revealed the isolation and identification of Pseudomonas flouresence from some moribund Oreochromis niloticus. Higher performance parameters were recorded in group 2 that received feeds treated with clay which reflected in the total production which reaches 1646.47 kg while in the control pond, the total production was1308.36 kg.

Molecular Cloning, Phylogenetic Analysis, Expressional Profiling and In Vitro Studies of TINY2 from Arabidopsis thaliana

  • Wei, Gang;Pan, Yi;Lei, Juan;Zhu, Yu-Xian
    • BMB Reports
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    • v.38 no.4
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    • pp.440-446
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    • 2005
  • A cDNA that was rapidly induced upon abscisic acid, cold, drought, mechanical wounding and to a lesser extent, by high salinity treatment, was isolated from Arabidopsis seedlings. It was classified as DREB subfamily member based on multiple sequence alignment and phylogenetic characterization. Since it encoded a protein with a typical ERF/AP2 DNA-binding domain and was closely related to the TINY gene, we named it TINY2. Gel retardation assay revealed that TINY2 was able to form a specific complex with the previously characterized DRE element while showed only residual affinity to the GCC box. When fused to the GAL4 DNA-binding domain, either full-length or its C-terminus functioned effectively as a trans-activator in the yeast one-hybrid assay while its N-terminus was completely inactive. Our data indicate that TINY2 could be a new member of the AP2/EREBP transcription factor family involved in activation of down-stream genes in response to environmental stress.

Modulation of the Cytochrome c Oxidase Activity by ATP: Implications for Mitochondrial Respiratory Control

  • Park, Nan-Hyang;Chun, Sun-Bum;Han, Tae-Young;Han, Sang-Hwa
    • BMB Reports
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    • v.29 no.4
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    • pp.300-307
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    • 1996
  • ATP and ADP are potential regulators of mitochondtial respiration and at physiological concentrations they affect the rate of electron transfer between cytochrome c and cytochrome c oxidase. The electron transfer, however, depends on the electrostatic interaction between the two proteins. In order to exclude any nonspecific ionic effects by these polyvalent nucleotides, we used 2'-O-(2,4,6)trinitro(TNP)-derivatives of ATP and ADP which have three orders of magnitude higher affinity for cytochrome c oxidase. A simple titration of the fluorescence intensity of TNP by cytochrome c oxidase showed a binding stoichiometry of 2:1 cytochrome c:cytochrome c oxidase. Higher ionic strength was required for TNP-ATP than for TNP-ADP to be dissociated from cytochrome c oxidase, indicating that the negative charges on the phosphate group are at least partially responsible for the binding. In both spectrophotometric and polarographic assays, addition of ATP (and ADP to a less extent) showed an enhanced cytochrome c oxidase activity. Both electron paramagnetic resonance and fluorescence spectra indicate that there is no Significant change in the cytochrome c-cytochrome c oxidase interaction. Instead, reduction levels of the cytochromes at steadystate suggest that the increased activity of nucleotide-bound cytochrome c oxidase is due to faster electron transfer from cytochrome ${\alpha}$ to cytochrome ${\alpha}_3$, which is known to be the fate limiting step in the oxygen reduction by cytochrome c oxidase.

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