• Title/Summary/Keyword: Gel chromatography

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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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Characterization of Vitamins in Yeast Extract using Gel Filtration, Ion Exchange Chromatography and HPLC (젤 여과, 이온 크로마토그래피와 HPLC에 의한 효모 엑기스내의 비타민의 분석연구)

  • 최인호;홍억기;강환구;김인호
    • KSBB Journal
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    • v.15 no.1
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    • pp.76-79
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    • 2000
  • Complex, ill-defined mixtures of natural origin are often used as nutrients in the production of biological products through microbial fermentation. Product yields are affected by variation in these natural products. Yeast extract is a typical example of these natural products. Since it is a mixture of amino acids, peptides and nucleic acids, its composition is not well characterized. In this study, we investigated the properties of thiamine hydrochloride, riboflavin and pyridoxine hydrochlride in yeast extract by using a gel filtration chromatography, ion exchange chromatography and high performance liquid chromatography. Yeast extract solution was fractionated by gel filtration chromatography and ion exchange chromatography, and then, each fraction was analyzed by using a high performance liquid chromatography.

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Subunits of Sweet Potato $\beta$-Amylase (고구마 $\beta$-아밀라제의 subunit)

  • 안용근
    • The Korean Journal of Food And Nutrition
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    • v.1 no.2
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    • pp.50-55
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    • 1988
  • B-Amylase was obtained from sweet potato extract in a crystalline state by dialysis against water after precipitated with acetone according to the method reported previously followed by DEAE Sephadex A-50 ion exchange chromatography plus gel chromatography of Sephadex G-200. The purified enzyme was homogeneous by SDS PAGE. The efforts had done to remove the miner bands in SDS PAGE by Sephadex G-200 gel chromatography, DATE Sephadex A-50 ion exchange chromatography. isoelectrophocusing, affinity chromatography, hydroxyapatite chromatography, recrystallizstion and HPLC on a column of TSK gel SW 3000 but have given any result. But, N -terminal amino acid of the enzyme was revealed mainly alanine and trace of glycine and glutamic acid. Therefore, it seems that the miner bands in SDS PAGE have a role of subunit.

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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 of 3,4-Dihydroxybenzoic Acid with Antimicrobial Activity from Bark of Aralia elata (두릅수피에서 항미생물활성을 갖는 3,4- dihydroxybenzoic acid의 분리)

  • Ma, Seung-Jin;Ko, Byoung-Seub;Park, Keun-Hyung
    • Korean Journal of Food Science and Technology
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    • v.27 no.5
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    • pp.807-812
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    • 1995
  • The methanol extracts of Aralia elata bark showed antimicrobial activities against bacteria, yeast, fungi. The solvent fractionated acidic fraction that showed the activity was then successively purified with silica gel adsorption column chromatography, Sephadex LH-20 column chromatography, silica gel partition column chromatography, HPLC and TLC. The isolated major active substance was identified as 3,4-dihydroxybenzoic acid by MS, GC-MS, IR, $^{1}H-NMR\;and\;^{13}C-NMR$. The content of 3,4-dihydroxybenzoic acid was 0.869㎎/g in dried bark of Aralia elata.

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Antimicrobial Substances in Leek (Allium tuberosum) (부추의 항미생물 활성물질)

  • Kim, Seon-Jae;Park, Keun-Hyung
    • Korean Journal of Food Science and Technology
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    • v.28 no.3
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    • pp.604-608
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    • 1996
  • The antimicrobial activty of leek (Allium tuberosum) was investigated against 17 strains of microorganisms. Methanol extracts of leek showed the growth inhibition effects on the wide range of microorganisms including gram positive bacteria, gram negative bacteria and yeasts. The extracts were analysed by using solvent fractionation, silica gel adsorption chromatography, Sephadex LH-20 column chromatography, TLC, silica gel partition chromatography and HPLC techniques. Six components whose molecular weights range from 200 to 400 were confirmed to have the antimicrobial activity.

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Brassinosteroid substances in immature Cassia tora seeds (결명자의 brassinosteroid 활성물질)

  • Park, Keun-Hyung;Kim, Seon-Jae;Hyun, Kyu-Hawn
    • Applied Biological Chemistry
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    • v.36 no.2
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    • pp.99-104
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    • 1993
  • In order to explore the brassinosteroid-active component in Cassia tora, methanol extract of immature seeds was purified by sequences of solvent fractionation, silica gel adsorption chromatography, Sephadex LH-20 chromatography, charcoal adsorption chromatography and Bondesil chromatography. The activity of brassinosteroid was monitored by the rice inclination test and its presence could be confirmed in each purification step. The purified active components were separated by silica gel adsorption chromatography. Brassinosteroid substances in separated active fractions were identified as teasterone, castasterone, brassinolide by TLC and HPLC. Our work is probably the first report of endogenous brassinosteroid in Cassia tora. The content of brassinosteroid in Cassia tora as converted into brassinolide was $3.5{\sim}5.5\;ng/g$ fresh weight. The order of brassinosteroid contents was toward to be teasterone, castasterone, brassinolide.

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Purification of $\beta$-Galactosidase from Alkalophilic Bacillus sp. YS-309 (호알카리성 Bacillus sp. YS-309로부터 $\beta$-Galactosidase의 정제)

  • 유주현;윤성식
    • Microbiology and Biotechnology Letters
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    • v.17 no.6
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    • pp.587-592
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    • 1989
  • A strain of alkalophilic Bacillus sp. YS-309 capable of producing large amount of $\beta$-galactosidase has been isolated from soil sample. Intracellular $\beta$-galactosidase was purified 6.9 folds by procedures including ammonium sulfate precipitation, DEAE-cellulose chromatography, gel-filtration, DEAE-Sephadex A-50 chromatography with over-all yield of 17.8%. The molecular weight of native enzyme was 205, 000 by HPLC, and SDS-polyacrylamide gel electrophoresis showed that the enzyme consisted of 4 identical subunits with a molecular weight of 56, 000.

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Purification of the Candida utilis Extracellular Invertase using Affinity Chromatography

  • Ginalska, G.;Belcarz, A.;Lobarzewski, J.;Leonowicz, A.;Cho, Nam-Seok
    • Journal of the Korean Wood Science and Technology
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    • v.30 no.3
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    • pp.12-17
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    • 2002
  • The extracellular invertase (EC 3.2.1.26) (Candida utilis) preparation was obtained from the liquid medium after desalting and freeze drying. This prepared enzyme was used for the comparative purification on 4 activated matrices by liquid column affinity chromatography method. In this method there were used controlled porous glass (CPG) silanized covalently activated by keratin, silanized silica gel and silica gel covalently covered by keratin. It was found that the invertase purification process was better using both CPG matrices (silanized CPG and keratin activated CPG) than these with two silica gel supports. Also the elution coefficient of the invertase from the two CPG columns was about 93 to 94%. Two silica gel supports found to be superior in terms of purification efficiency. The invertase purification process was confirmed by PAGE electrophoresis.