• Title, Summary, Keyword: gel filtration

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Purification of Peptide Components including Melittin from Bee Venom using gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis (Gel filtration chromatography와 propionic acid/urea polyacrylamide gel electrophoresis를 이용한 봉독 성분의 분리)

  • Choi, Young-Chon;Choi, Suk-Ho;Kwon, Ki-Rok
    • Journal of Pharmacopuncture
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    • v.9 no.2
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    • pp.105-111
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    • 2006
  • Objectives : This study was conducted to carry out Purification of Melittin and other peptide components from Bee Venom using gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis Methods : Melittin and other peptide components were separated from bee venom by using gel filtration chromatography on Sephadex G-50 column in 0.05M ammonium acetate buffer. Results : Melittin and other peptide components were separated from bee venom by using gel filtration chromatography on Sephadex G-50 column in 0.05M ammonium acetate buffer. The fractions obtained from gel filtration chromatography was analyzed by using SDS-PAGE and propionic acid/urea polyacrylamide gel electrophoresis. The melittin obtained from the gel filtration contained residual amount of phospholipase $A_2$ and a protein with molecular weight of 6,000. The contaminating proteins were removed by the second gel filtration chromatography. Conclusion : Gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis are useful to separate peptide components including melittin from bee venom.

Purification and Characterization of a Juvenile Hormong Binding Protein from Whole Body Homogenates of the Wax Moth, Galleris mellonella Final Instar Larvae (꿀벌부채명나방 종령유충에서 유약호르몬 결합단백질의 정제와 특성)

  • 안기흥;전상학;이경로
    • Korean journal of applied entomology
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    • v.37 no.1
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    • pp.59-64
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    • 1998
  • A juvenile hormone binding protein (JHBP) has been isolated from the whole body homogenate of Galleria rnellonella final instar larvae by gel filtration. The isolated protein is homogenous as judged by column chromatography and gel electrophoresis in the presence and absence of denaturing agent. The JHBP has a relative molecular weight of 32 k by denaturing gel electrophresis and 28 k by gel filtration. The protein exhibits a dissociation constant of 3.9 x M for JH 111.

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Protein-silica Interaction in Silica-based Gel Filtration Chromatography (Silica-based Gel Filtration 크로마토그래피에서의 단백질-실리카 상호작용)

  • Choi, Jung-Kap;Yoo, Gyurng-Soo
    • YAKHAK HOEJI
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    • v.35 no.6
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    • pp.461-465
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    • 1991
  • Silica-based gel filtration chromatography has been used to characterize molecular weight of proteins. However, the molecular weight measured by this method was distorted by protein-silica interactions like hydrophobic and electrostatic forces. Therefore, we characterized protein-silica interaction using two forms of phytochrome (124 kDa) having different hydrophobicity and surface charge. PH and ionic strength affected the retention time of phytochrome suggesting that electrostatic force is the major interaction between protein and silica surface.

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The Properties of Acetolactate Synthase Isozyme Produced by Serratia marcescens ATCC 254 19 (Serratia marcescens ATCC 25419가 생산하는 Acetolactate Synthase Isozyme의 특성)

  • 김종탁;김승수
    • Microbiology and Biotechnology Letters
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    • v.20 no.1
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    • pp.25-33
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    • 1992
  • One acetolactate synthase isozyme which has Rf value of 0.83 on polyacrylamide gel electrophoresis was purified from Sewatia marcescens ATCC 25419 by ammonium sulfate fractionation, DEAE-Sephacel chromatography, Phenyl-Sepharose chromatography, Sephacryt S-400 gel filtration followed by native gel elution. The native molecular weight of the enzyme was determined to be 531,400 by gel filtration method, and SDS-polyacrylamide gel electrophoresis separated the native enzyme into two polypeptides having molecular sizes of 55,000 and 38,900 respectively. In kinetic parameters, $K_m$ value for pyruvate was 2.54 mM, and $V_{max}$ was 21.75 nmoie/min/mg. The enzyme showed maximal activity around pH 8.0 and optimal temperature of the acetolactate formation was $37^{\circ}C$. Feedback inhibition studies indicate that the purified enzyme is rather resistant to branched chain amino acids when compared with acetolactate synthase isozymes of plants or other enterobacteria.

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Studies on the Isolation and Immunochemical Properties of SIgA from Human and Bovine Milk (인유(人乳) 및 우유(牛乳)로부터 Secretory Immunoglobulin A의 분리(分離) 및 면역화학적(免疫化學的) 특성(特性)에 관(關)한 연구(硏究))

  • Lee, Jo Yoon;Kim, Jong Woo
    • Korean Journal of Agricultural Science
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    • v.22 no.1
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    • pp.82-95
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    • 1995
  • These experiments were carried out to isolate SIgA from human and bovine milk. The immunochemical properties of SIgA from human and bovine milk were examined by Gel filtration, DEAE and SDS-PAGE. Double Immunodiffusion, and Immunoelectrophoresis. The results obtained are as follows: 1. Human SIgA was purified from colostrum of Korean women by repeated gel filtration on Sephadex G-200 and Sepharose 6B, but bovine SigA was not cleary purified from bovine colostrum of Holstein cows by anion exchange chromatography using DEAE-Sephadex A-50 and gel filtration on Sephadex G-200 and Sepharose 6B. 2. The immunochemical properties of fractions from gel filtration on the Sephadex G-200 and Sepharose 6B column as assessed by Immunoelectrophoresis and double Immunodiffusion to identify the presence of IgM in first peak fraction, and the presence of pure SIgA in second peak fraction. However, Bovine SigA rich fraction from bovine colostrum of Holstein cows contained a large amount of $IgG_1$-dimer in addition to SIgA. 3. The fragments of reduced bovine colostrum SIgA rich fraction were estimated to have molecular weights of secretory component, heavy chain and light chain (75,000-80,000, 50,000-60,000, 25,000-27,000 daltons) by SDS-PAGE, respectively. Those were similar to the molecular weight of reduced SIgA from human milk.

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Purification and Properties of Isocitrate Lyase from Saccharomycopsis lipolytica (Saccharomycopsis lipolytica Isocitrate Lyase의 정제와 성질)

  • 조석금
    • Microbiology and Biotechnology Letters
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    • v.15 no.6
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    • pp.420-424
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    • 1987
  • Isocitrate lyase from crude extract of Saccharomycopsis lipolytica ATCC44601 and MX9-11RX8 temperature-sensitive mutant was purified about 54 times and 87 times, respectively by ammonium sulfate fractionation, Toyo peal HW-55F gel filtration and DEAE-Cellulose ion exchange chromatography, The molecular weight of the purified isocitrate lyase from this yeast was estimated to be 230, 000 by gel filtration on Sephadex G-200, and SDS-polyacrylamide Eel electrophoresis showed that the enzyme consisted of four identical or similar subunits with a molecular weight of 59, 000 and the enzyme showed optimum activity at pH 6.9.

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Study on the Purification of Transforming Growth Factor-$\beta$ in Canine Platelets (개 혈소판에서 변형성장인자 베타의 분리에 관한 연구)

  • Kweon Oh-Kyeong;Hong Sung-Hyeok
    • Journal of Veterinary Clinics
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    • v.11 no.1
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    • pp.389-392
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    • 1994
  • To purify transforming growth factor type beta(TGF-$\beta$) in canine platelets, Sephadex G-75 gel filtration and semipreparative HPLC were carried out. The column of $2.0 {\times}120cm$ was used for gel filtration and one inch semipreparative column filled with SP-Toyopeal for HPLC. Electrophoresis and bioassay using African green monkey kidney cell were used for identification of TGF-$\beta$ Crude TGF-$\beta$ of 2.75mg was extracted from 5.2g of the platelets by the treatment of acid/ethanol. In gel filtration of crude TGF-$\beta$, 4 peaks were observed at the detection of spectrophotometer at 280nm. Electrophoresis and bioassay identified the 3rd peak TGF-$\beta$. Linear gradient elution from 0 to 3M NaCl in sornipreparative HPLC showed TGF-$\beta$ at 1.5M NaCl. Gel filtration was less expensive and useful method for the purification of TGF-$\beta$.

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Identification and Isolation of Juvenile Hormone Binding Protein from Hemolyrnph of Lymantria dispar L. (매미나방(Lymantria dispar)에서 Juvenile Hormone Binding Protein(JHBP)의 확인 및 정체)

  • 이인희;김학열
    • The Korean Journal of Zoology
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    • v.34 no.2
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    • pp.196-202
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    • 1991
  • Juvenile honnone binding protein (JHBP) was identified in the last instar larval hemolymph of Lymantria dispar using dextran coated charcoal (DCC) binding assay and gel filtration. The p1 value of JHBP was estimated to be 5.3. JHBP was partially pudfied by polyethylene glycol(PEG) precipitation, DEAE-cellulose ion-exchange chromatography and gel filtration, and was confirmed by DCC binding assay.

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Pure-Separation of Calcium chloride-treated Silk Fibroin Hydrolysate by Gel Filtration Chromatography and Effect of It's Enzymatic Hydrolysis (Calcium chloride 피브로인 용해물의 Gel Filtration Chromatography에 의한 순수분리 및 효소 가수분해 효과)

  • 여주홍;이광길;이용우
    • Journal of Sericultural and Entomological Science
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    • v.41 no.3
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    • pp.211-215
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    • 1999
  • The pure-separation of calcium chloride-treated fibroin hydrolysates could be carried out using gel filtration chromatography. Also, the effect of its enzymatic hydrolysis was investigated in order to find out the enhancement of their functionality. The average molecular weight(Mw), solubility and free amino acid compositions of three hydrolysates samples (calcium chloride, calcium chloride-flavourzyme and calcium chloride-thermoase)were measured to compare their characteristics. The molecular weight of calcium chloride hydrolysate was about Mw 46,800 and it can be reduced to Mw 12,500 and 1,070 upon the enzymatic hydrolysis by flavourzyme and thermoase, repectively. A solubility of calcium chloride-treated samples shows about 60% while calcium chloride/enzyme-treated samples are perfectly soluble (100% solubility). The total amino acid composition of calcium chloride enzymatic hydrolysates are much higher than that of calcium chloride hydrolysate.

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Isolation and Characterization of a Nematicidal Bacillus thuringiensis strain 108 (항선충성 Bacillus thuringiensis 108균주의 분리와 특성)

  • Lee, Jae-Hun;Ryu, Eun-Ju;Kim, Kwang-Hyeon
    • Microbiology and Biotechnology Letters
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    • v.35 no.3
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    • pp.250-254
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    • 2007
  • Bacillus thringiensis strain 108 was isolated from soil and had nematicidal activity against second stage juvenile of plant root-knot nematode, Meloidogyne incognita. The strain 108, a rod shape, spore forming and Gram positive bacterium, produced lecithinase, catalase, and ${\delta}$-endotoxin. The strain 108 belongs to H serotype 3, Bacillus thuringiensis var. kurstaki. A nematicidal substance of the strain 108 was partially purified on Sephadex G-25 gel filtration, activated carbon adsorption, silica gel adsorption, and Sephadex G-10 gel filtration. $LC_{90}$ of the partially purified substance against M incognita was $1.2\;{\mu}g/ml$. The nematicidal substance was stable by heat treatment at $100^{\circ}C$ for 1hr, but was perfectly lost nematicidal activity after autoclave ($110^{\circ}C$, 30 min).