• Title/Summary/Keyword: hemagglutinating activity

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Overexpression and Characterization of Vibrio mimicus Metalloprotease

  • Shin, Seung-Yeol;Lee, Jong-Hee;Huh, Sung-Hoi;Park, Young-Seo;Kim, Jin-Man;Kong, In-Soo
    • Journal of Microbiology and Biotechnology
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    • v.10 no.5
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    • pp.612-619
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    • 2000
  • To investigate the biochemical properties of V. mimicus metalloprotease, whose gene was isolated previously from Vibrio mimicus ATCC33653, overexpression and purification were attempted. The 1.9 kb of open reading frame was amplified by PCR from pVMC193 plasmid which ligated the VMC gene with pUC19 and introduced into Escherichia coli BL21 (DE3) using the overexpression vector, pET22b (+). The overexpressed metalloprotease (VMC) was purified with Ni-NTA column chromatography and characterized with various protease inhibitors, pHs, temperatures, and substrates. The purified VMC showed the proteolytic activity against gelatin, soluble and insoluble collagens, and synthetic peptides. Unlike the observations made with all metalloproteases originated from other Vibrio sp., the VMC did not hydrolyze the casein. The proteolytic activity was critically decreased when the VMC was treated with metal chelating reagents, such as EDTA, 2,2-bipyridine, and 1, 10-phenanthroline. In particular, the 71 kDa VMC exhibited the hemagglutinating activity against human erythrocyte. As the purified VMC was treated with $CuCl_2$ and $NiCl_2$ for the chemical modification of metal binding, the proteolytic activity and hemagglutinating activity were profoundly influenced. The multialignment analysis made on the reported Vibrio metalloproteases showed the difference of amino acid sequence similarity between the two distinctive classes of Vibrio metalloproteases.

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Isolation and Characterization of a Trypsin Inhibitor and a Lectin from Glycine max cv. Large Black Soybean

  • Ye, Xiu Juan;Ng, Tzi Bun
    • Food Science and Biotechnology
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    • v.18 no.5
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    • pp.1173-1179
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    • 2009
  • Trypsin inhibitors and lectins are defense proteins produced by many organisms. From Chinese 'Large Black Soybeans', a 60 kDa lectin and a 20 Da trypsin inhibitor (TI) were isolated using chromatography on Q-Sepharose, Mono Q, and Superdex 75. The TI inhibited trypsin and chymotrypsin with an $IC_{50}$ of 5.7 and $5{\mu}M$, respectively. Trypsin inhibitory activity of the TI was stable from pH 3 to 13 and from 0 to $65^{\circ}C$. Hemagglutinating activity of the lectin was stable from pH 2 to 13 and from 0 to $65^{\circ}C$. The TI was inhibited by dithiothreitol, signifying the importance of disulfide bond. The TI and the lectin inhibited HIV-1 reverse transcriptase ($IC_{50}$=44 and $26{\mu}M$), and proliferation of breast cancer cells ($IC_{50}$=42 and $13.5{\mu}M$) and hepatoma cells ($IC_{50}$=96 and $175{\mu}M$). The hemagglutinating activity of the lectin was inhibited most potently by L-arabinose. Neither the lectin nor the TI displayed antifungal activity.

Characterization of the recombinant metalloprotease from Vibro mimicus and its hemagglutinating activity

  • Kong, In-Soo;Shin, Seung-Yeol;Lee, Jong-Hee;Kim, Jin-Man;Park, Young-Seo
    • Proceedings of the Korean Society of Fisheries Technology Conference
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    • pp.161-162
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    • 2000
  • Metalloprotease produced in Vibrio mimicus, in which zinc is an essential meta ion for catalytic activity, degrades a variety of biologically important substances including human collagen, several complement components, and immunoglobulin. For gene overexpression and convenient purification, VMC gene was constructed in pET22b(+) expression vector by using of PCR. (omitted)

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Production and characterization of monoclonal antibody against bovine coronavirus (소 코로나바이러스에 대한 단크론항체 생산과 특성)

  • Ahn, Jae-moon;Kang, Shien-young
    • Korean Journal of Veterinary Research
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    • v.38 no.3
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    • pp.581-588
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    • 1998
  • Eight monoclonal antibodies(MAbs) against bovine coronavirus(BCV) were produced and characterized. Three MAbs(1G9, 4H12, 5C1) specific to the S glycoprotein and two HE glycoprotein-specific MAbs(2A5, 5G4) were found to neutralize the BCV in fluorescence focus neutralization(FFN) test. Two HE-specific MAbs from the neutralizing MAbs inhibited the hemagglutinating activity of the BCV. None of the N protein-specific MAbs(1C1, 5A12, 6H1) neutralized the virus infectivity. Bovine coronavirus and mouse hepatitis virus, which belong to group II coronaviruses, were differentiated from other groups of coronaviruses(porcine transmissible gastroenteritis virus, porcine epidemic diarrhea virus, canine coronavirus) by all MAbs in fluorescence antibody test(FA), but not in FFN test.

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Comparison of Lectin from Pseudixus japonicus and Concanavalin a on Lymphocytes Proliferation and Cytotoxicity

  • Chung, Yong-Za;Jung, Hyun-Ok;Hong, Tae-Hong;Suh, Sok-Soo
    • Archives of Pharmacal Research
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    • v.14 no.3
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    • pp.207-216
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    • 1991
  • Pseudixus japonicus agglutinin (PJA) was isolated. And its characteristics were compared with those of concanavalin A (Con A). PJA is a glycopritein composed of 49.3% carbohydrate and 50.7% protein which had relatively high percentages of glutamic acid, aspartic acid and phenylalanine residues. The hemagglutinating activity of PJA was approximately one-eighth of that of Con A when tested with mouse crythrocytes. PJA failed to simulate the proliferation or transformation of human and mouse lymphocytes in contratst to Con A. PJA and Con A showed cytotoxicities against SNU-1 (human stomach cancer cells), SNU-CI (human colon cancer cells) and mouse Sarcoma 180 cells when tested by 3-(4, 5-dimethyl thiazol-2-yl)2. 5-diphenyl tetrazolium bromide (MIT) colorimetric assay. The antitumor activity of the lectin in vivo was also tested in Sarcoma 180 bearing mice. There was no significant difference in prologation of lifc span of the mice after the treatment with PJA and Con A for 10 consecutive days.

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Functional Display of Maackia amurensis Hemagglutinin (MAH) on Bacteriophage (박테리오파아지 표면 발현 시스템을 이용한 Maackia amurensis Hemagglutinin (MAH)의 기능적 발현)

  • 임미정
    • YAKHAK HOEJI
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    • v.47 no.3
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    • pp.176-179
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    • 2003
  • A library of unlimited number of novel lectins with diverse specificities has been previously generated by randomly mutating the carbohydrate-recognition domain of Maackia amurensis hemagglutinin (MAH). To establish the experimental environment capable of selecting high affinity mutant lectins in E. coli, phage display system was adapted. Carbohydrate binding capacity of two phagemid vectors, pComb3 and pComb8 displaying wild-type MAH lectin was assessed. Specific bindings of pComb3 and pComb8 phages expressing w.t. MAH to affinity-purified polyclonal anti-MAH antibody and to glycophorin was demonstrated. Both phages also showed strong hemagglutinating activity to intact but not sialidase-treated human erythrocytes, which is consistent to the specificity of native MAH. Taken together, two different phage display vectors successfully allowed the expression of active MAH as a fusion protein on the surface of bacteriophage, which will lead to preparation of unique plant lectins with high affinity toward a variety of carbohydrate chains.

Trypsin Inhibitor and Hemagglutinating Activities of Some Minor Beans in Korea (한국산 두류(頭類)의 Trypsin 저해(沮害) 활성(活性) 및 적혈구(赤血球) 응집(凝集) 활성(活性))

  • Kang, Myung-Hee;Kim, Yong-Hwa;Lee, Su-Rae
    • Korean Journal of Food Science and Technology
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    • v.12 no.1
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    • pp.24-33
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    • 1980
  • Trypsin inhibitor and hemagglutinating activities of some minor beans produced in Korea were determined in comparison with those of soybean and the effects of heat treatment on the activities were studied. The results are summarized as follows 1. The trypsin inhibitor activity (% [TU] inhibited/mg) of soybean, red bean, kidney bean, and mung bean were 79.9, 46.4, 43.2 and 17.7, respectively, on the dry weight basis and were 194, 222, 170 and 75, respectively, on the protein basis. 2. Heat destruction by boiling or autoclaving of trypsin inhibitor activity of red bean, mung bean, kidney bean, and soybean were $85{\sim}87,\;87{\sim}94,\;76{\sim}79\;and\;67{\sim}72\;%$, respectively. No significant difference was, however, observed in the effect between the two heating methods. 3. The hemagglutinating activity (unit/g) of kidney bean, soybean, mung bean and red bean were 48,300, 18,000, 136 and non-detectable, respectively, on the dry weight basis and were 190,600, 43,700, 581 and non-detectable, respectively, on the protein basis. Heat treatment destructed the hemagglutinating activity in all three beans.

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Purification and Characterization of Hemagglutinating Protein from Rhizome of Alisma orientale (택사(Alismatis Rhizoma) Hemagglutinating Protein의 정제와 특성)

  • 박종옥;김경순;선우근옥
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.24 no.4
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    • pp.587-593
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    • 1995
  • Lectin was purified by using $(NH_4)_2SO_4$, DEAE-cellulose ion-exchange chromatography and Sephadex G-150 column chromatography from Alismatis Rhizoma(AR). The specific activity of AR lectin was 50, 441units/mg, and purification folds were 114. The AR lectin agglutinated human erythrocytes of all types(A, B, O, AB). The molecular weight of AR lectin was estimated about 90, 500 daltons by gel filtration and each subunits were 42,000, 27,000 and 22,500 daltons on SDS-PAGE respectively. The hemagglutinating activity of the lectin was inhibited by sialic acid, glucose, ribose, galactose, sucrose, and lactose. It was also inhibited by cations such as $Hg^{++},\;Fe^{++},\;Cu^{++}\;and\;Pb^{++}$.

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Isolation and Characterization of Lectins from Stem and Leaves of Korean Mistletoe (Viscum album var. coloratum) by Affinity Chromatography

  • Park, Won-Bong;Han, Seon-Kyu;Lee, Myung-Hwang;Han, Kwang-Ho
    • Archives of Pharmacal Research
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    • v.20 no.4
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    • pp.306-312
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    • 1997
  • We attempted to isolate and characterize the lectins from stem and leaves of Korean mistletoe (Viscum album var. coloratum) by affinity chromatography. Lectin I was isolated only from stem. Lectin II was not isolated from Korean mistletoe, whereas lectin III was isolated from the stem and leaves. The hemagglutinating activity of lectin I was 16HU and inhibited by D-galactose, lactose, and N-acetyl-D-galactosamine. The lectin I has molecular weight of 60, 000D being composed of two basic subunits with molecular weights of 32, 000D and 28, 000D which are linked by a disufide bond. The lectin III from stem has molecular weight of 66, 000D being two basic subunits which have molecular weights of 34, 000D and 29, 000D and are linked by a disufide bond. The activity of lectin I was stable at the pH range of 4.00-8.50 and at a wide range of temperature (0-42.deg. C). The lectin I showed more potent mitogenic activity to murine lymphocytes than concanavalin A.

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A Lectin with Mycelia Differentiation and Antiphytovirus Activities from the Edible Mushroom Agrocybe aegerita

  • Sun, Hui;Zhao, Chen Guang;Tong, Xin;Qi, Yi Peng
    • BMB Reports
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    • v.36 no.2
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    • pp.214-222
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    • 2003
  • A lectin named AAL has been purified from the fruiting bodies of the edible mushroom Agrocybe aegerita. AAL consisted of two identical subunits of 15.8 kDa, its pI was about 3.8 determined by isoelectric focusing, and no carbohydrate was discerned. Being treated by pyrogultamate aminopeptidase, the blocked N-terminus of AAL was sequenced as QGVNIYNI. AAL agglutinated human and animal erythrocytes regardless of blood type or animal species. Its hemagglutinating activity was unaffected by acid or alkali treatment and demetalization or addition of divalent metals $Mg^{2+}$, $Ca^{2+}$ and $Zn^{2+}$. AAL was toxic to mice: its LD50 was 15.85 mg per kilogram body weight by intraperitoneal injection. In this study, two novel activities of AAL were proved. It showed inhibition activity to infection of tobacco mosaic virus on Nicotiana glutinosa. The result of IEF suggested that AAL attached to TMV particles. Mycelia differentiation promotion was the other interesting activity. AAL promoted the differentiation of fruit body primordia from the mycelia of Agrocybe aegerita and Auricularia polytricha. AAL antiserum was prepared and immunologically cross-reactived with several proteins from five other kinds of mushrooms. These results suggested that AAL probably was a representative of a large protein family, which plays important physiological roles in mushroom.