Relevant bibliographies by topics / Gel filtration chromatography

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  2. Dissertations / Theses
  3. Books
  4. Book chapters
  5. Conference papers

Academic literature on the topic 'Gel filtration chromatography'

Author: Grafiati
Published: 4 June 2021
Last updated: 20 February 2023

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Journal articles on the topic "Gel filtration chromatography"

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Werner, Milton H., G. Marius Clore, Angela M. Gronenborn, Akiko Kondoh, and Robert J. Fisher. "Refolding proteins by gel filtration chromatography." FEBS Letters 345, no. 2-3 (May 30, 1994): 125–30. http://dx.doi.org/10.1016/0014-5793(94)00401-3.

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Barnes, Ana Isabel, Cristina Ortiz, María Gabriela Paraje, Luis Eduardo Balanzino, and Inès Albesa. "Purification and characterization of a cytotoxin fromEnterobacter cloacae." Canadian Journal of Microbiology 43, no. 8 (August 1, 1997): 729–33. http://dx.doi.org/10.1139/m97-105.

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Leukotoxic activity was assayed in clinical isolates of Enterobacter cloacae. Two strains were selected out of 38 by their greater hemolytic activity in blood agar plates. Leukotoxin was purified by salt precipitation, dialysis, chromatography by gel filtration, and high pressure liquid chromatography (HPLC). Human leukocytes, when incubated with purified E. cloacae toxin, showed high percentages of death and lysis, with time and dose dependence. The chromatographic profile of gel filtration presented three protein peaks and toxic activity was detected in the second peak. After HPLC, leukotoxin coeluted with the hemolytic activity and both activities were detected only after 2-mercaptoethanol treatment. Coomassie-stained sodium dodecyl sulfate – polyarylamide gels showed a single band. This band was estimated to represent a protein of 13 300 Da on the basis of both sodium dodecyl sulfate – polyacrylamide gel electrophoresis and gel filtration chromatography.Key words: Enterobacter clocae, leukotoxin, molecular mass, cytotoxin, leukocytes.
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Hunt, Eric A., and Sapna K. Deo. "Board-Game Gel Filtration and Affinity Chromatography." Journal of Chemical Education 86, no. 1 (January 2009): 19. http://dx.doi.org/10.1021/ed086p19.

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Wooten, Arthur L., M. Lynn Prewitt, Terry Sellers, and David C. Teller. "Gel filtration chromatography of resole phenolic resins." Journal of Chromatography A 445 (January 1988): 371–76. http://dx.doi.org/10.1016/s0021-9673(01)84549-0.

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Girona, V., J. Estelrich, M. Pujol, and J. Bolòs. "Ampicillin polymers: identification by gel-filtration chromatography." International Journal of Pharmaceutics 41, no. 3 (February 1988): 241–44. http://dx.doi.org/10.1016/0378-5173(88)90200-1.

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Kumar Shaha, Ranajit, Nitai Roy, and Talukdar G. "Characterization and Sensitivity Test of the Allergenic Pollen Proteins from Litchi Chimensis Plant." Journal of Tropical Resources and Sustainable Science (JTRSS) 1, no. 1 (August 15, 2021): 25–35. http://dx.doi.org/10.47253/jtrss.v1i1.667.

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Pollen of Litchi chinensis (Litchi) is a major aeroallergen of Bangladesh. Pollen of this fruits plant was collected from full bloomed flower growing in different places of Rajshahi in Bangladesh. Pollen protein was extracted and partial purified by means of long-term PBS extraction, salting out, dialysis, gel filtrations and DEAE-Cellulose chromatography and the protein was designated as LFPP (Litchi flowers pollen protein). Gel filtration of the purified pollen protein gives two main peaks. The major peak gives four bands on SDS-PAGE. The enzyme (pectate lyase) proteins after gel filtration again re-purified by Ion exchange chromatography, a single band in the protein profile of LFPP, (M.W. 28kDa) was the major allergenic component of Litchi chinensis (Litchi) flower pollen. The homogeneity and the molecular weight of the protein were estimated by SDS-PAGE, and Gel filtration was 28kDa. The allergenic protein was identified by skin prick tests and showed the pectate lyase (Pel) activity. Skin-prick tests also revealed highest degree of sensitivity to the Nawabgang sample giving positive response in 80% of the patients. Skin reactivity ranged between 1+ and 3+.
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Russo, Salvatore F., and Angie Radcliffe. "Separations utilizing gel filtration and ion-exchange chromatography." Journal of Chemical Education 68, no. 2 (February 1991): 168. http://dx.doi.org/10.1021/ed068p168.

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González, Ana, and Jaime Gómez-Márquez. "Purification of bacteriophage DNA by gel filtration chromatography." Gene Analysis Techniques 7, no. 1 (February 1990): 2–4. http://dx.doi.org/10.1016/0735-0651(90)90037-g.

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Whisenant, E. C., B. K. A. Rasheed, and Y. M. Bhatnagar. "Plasmid purification using high-performance gel filtration chromatography." Nucleic Acids Research 16, no. 11 (1988): 5202. http://dx.doi.org/10.1093/nar/16.11.5202.

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Nitoda, Teruhiko, Hirokazu Usuki, and Hiroshi Kanzaki. "A Potent Insect Chitinase Inhibitor of Fungal Origin." Zeitschrift für Naturforschung C 58, no. 11-12 (December 1, 2003): 891–94. http://dx.doi.org/10.1515/znc-2003-11-1226.

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Abstract A water-soluble polysaccharide was isolated from the culture filtrate of a fungal strain, Sphaeropsis sp. TNPT116-Cz, as a novel insect chitinase inhibitor. It was purified to chromatographic homogeneity by ethanol precipitation, anion-exchange and gel filtration chromatography. Its molecular weight was estimated to be 16 kDa by gel filtration HPLC. Monosaccharide analysis showed that it contained glucose, galactose, N-acetylglucosamine and a deoxysugar. This polysaccharide showed potent and specific inhibitory activity against Spodoptera litura chitinase with an IC50 value of 28 nᴍ.
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Dissertations / Theses on the topic "Gel filtration chromatography"

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Zhang, Qunying. "Characterization of receptors for Escherichia coli 987P using competitive binding assays, thin-layer chromatography and gel filtration chromatography." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0023/MQ51825.pdf.

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Ring, Ludwig. "Purification of psychoactive biomolecules in plants using size exclusion chromatography." Thesis, Linköping University, Department of Physics, Chemistry and Biology, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-18434.

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Size exclusion chromatography (SEC) was applied for purification of psychoactive biomolecules from plants. These molecules are in the same molecular weight range, but do not necessarily share other chemical properties, that makes the SEC technique efficient. By applying SEC as a first purification step much of the co-extractives from the plants can easily be removed. Large amounts of target substance can be obtained with little effort if the system is automated. Combining SEC with a second purification step, consisting of normal phase chromatography, provides high purity of the target substance.

Both known and unknown psychoactive biomolecules can easily be purified using the purification method developed in this Master's Thesis. Purifications that previously required long time and much "hands-on" can be completed much faster and with less manual work.

The method developed was tested on cannabis, coffee and 'Spice' with good results.

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Gustafsson, Sofia. "Expression and Purification of Murine Tripeptidyl Peptidase II." Thesis, Uppsala universitet, Institutionen för medicinsk biokemi och mikrobiologi, 2012. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-177009.

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Tripeptidyl peptidase II (TPPII) is an exopeptidase which cleaves tripeptides from theN-terminus of peptides. The exact functional role of TPPII is still a matter of investigation. Itis believed that the enzyme is primarily involved in intracellular protein degradation, where itcooperates with the proteasome and other peptidases to degrade proteins into free aminoacids. These amino acids can subsequently be used in the production of new proteins. The aimof this work was to express murine wild type TPPII using E. coli and thereafter purify theenzyme from the bacterial lysate. Methods used for the purification included protein andnucleic acid precipitation, anion exchange chromatography, hydrophobic interactionchromatography and gel filtration. The presence of TPPII was determined using activityassay, western blot and SDS-PAGE. Despite the fact that some modification is still needed,the purification yielded a total of 34μg TPPII with a purity of approximately 60%. Thispurified enzyme can be used for future functional characterization.
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Suttisansanee, Uthaiwan. "Biochemistry in Bacterioferritin." Thesis, University of Waterloo, 2006. http://hdl.handle.net/10012/2983.

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Bacterioferritin, an iron storage protein having a 24-subunit quaternary structure, was used as a model for the study of host-guest interactions and guest encapsulation, making use of its spherical cage-like structure. A hexahistidine-affinity tag fused to the C-terminus of each bacterioferritin subunit was constructed. The C-terminus of each subunit points toward the inside of the cavity, while the N-terminus is exposed on the surface of the protein. The hexaHistag was able to form strong interactions with a nickel-nitrilotriacetic acid linked dye molecule (guest) and this interaction was used in attempts to develop a principle to control guest molecule encapsulation within the spherical cavity of the 24-mer bacterioferritin protein molecule. The procedure involved (1) subunit dissociation under acidic pH, (2) affinity controlled dye-Histag binding with exposed C-terminal hexahistidine residues and (3) reassociation of the subunits at neutral pH. The encapsulation conditions involving step 1 and 3 were studied preliminarily using laser light scattering to measure size (hydrodynamic radius) of the protein particle with apoferritin as a model system as it resembles the size and structure of bacterioferritin. In order to encapsulate guest molecules, the emptied shell of bacterioferritin was generated by site-directed mutagenesis resulting in ferroxidase- as well as heme-free bacterioferritin mutants (E18A/M52L/E94A), and these mutants were used to examine protein stability before conducting encapsulation experiments. However, wild-type bacterioferritin possessed highest stability in maintaining its multisubunit structure; hence, it was used for the encapsulation studies. It was found that 100% bacterioferritin with hexahistidine tag at the C-terminus, and a combination of 60% bacterioferritin with hexahistidine tag at the C-terminus and 40% bacterioferritin without hexahistidine tag at the C-terminus yielded similar amounts of encapsulated guest molecules. This suggested that all hexahistidine at the C-terminus were not equally available for dye molecule binding.
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Vergnolle, Chantal. "I: purification et caracterisation de proteines de transfert de phospholipides, a partir de feuilles d'epinard (spinacia oleracea l. ). Ii: synthese in vitro des proteines vegetales : methodologie." Paris 6, 1986. http://www.theses.fr/1986PA066254.

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Des proteines, capables de faciliter des mouvements intermembranaires de phospholipides, ont ete isolees a partir de feuilles d'epinard. Ces proteines, appelees proteines de transfert de phospholipides, ont ete purifiees par les techniques classiques de chromatographie (filtration sur gel, echangeurs d'ions) ou par les techniques plus resolutives et plus rapides de chromatographie liquide a haute performance (colonnes echangeuses d'ions ou en phase inverse). Nous avons verifie la purete des fractions par electrophorese en presence de sds
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Trevisoli, Edilaine Della Valentina Gonçalves. "Purificação de eliciadores de defesa vegetal em soja e feijoeiro a partir de nematoides fitopatogênicos." Universidade Estadual do Oeste do Paraná, 2016. http://tede.unioeste.br:8080/tede/handle/tede/1475.

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The induction of resistance in plants to pathogens is an alternative method of disease control, wich involves activation of plant resistance mechanisms such as induction of phytoalexins. The elicitors molecules are able to induce and activate those responses, and therefore, techniques have sought to isolate and characterize fractions with elicitor character. The study aimed to purify, through ion exchange chromatography and gel filtration chromatography, eliciting molecules from pathogenic nematodes, and test them in phaseolin induction in beans hypocotyls beans and gliceolin in soybean cotyledons. The buffer solution Tris HCl 0.05 M (pH 6.8) was used as control and the acibenzolar-S-methyl (50 mg a.i. L-1) and Saccharomyces cerevisiae (20 mg mL-1) were used as induction standard treatments. Ion exchange chromatography (IEC) and gel filtration chromatography (GC) were performed to separate fractions with eliciting power from 500 female nematodes of Meloidogyne incognita and Meloidogyne javanica. For purification of elicitors from Meloidogyne javanica, through IEC, six glycidic fractions and six glycoproteins were obtained. These were purified on GC, obtaining sixty-three fractions. They have been classified according to their nature, as twenty-six glycidic and thirty-seven glycoprotein with molecular weights ranging from 29.19 to 2989.25 kDa. Regarding the elicitors purification of Meloidogyne incognita through IEC, nine glycidic and five glycoprotein fractions were obtained. From these fractions, a total of fifty-eight fractions was obtained through GC, twenty-five glycidic and thirty-three glycoprotein with molecular weights ranging from 37.42 to 200.32 kDa. From the fractions purified from Meloidogyne javanica eight had inducing potential of phaseolin. For gliceolin fifteen fractions showed inducing effect. Regarding the fractions purified from Meloidogyne incognita, no fraction has inductive potential of phaseolin superior to the standard treatment. However, twenty-two fractions suppressed phytoalexin inducing activity. For gliceolin ten fractions induced the same, whereas, twenty-three fractions suppressed the induction of gliceolin. Chromatography was efficient in the purification of elicitors compounds. Compounds with suppressing characteristics of gliceolin and phaseolin were checked in bioassays. For those fractions obtained through IEC, and then submitted to GC that did not induce phytoalexin, it is suggested that molecules need to act together to have elicitor effect and thus induce defense response in the plant
A indução de resistência em plantas contra patógenos é um método de controle alternativo de doenças, e que envolve a ativação dos mecanismos de resistência da planta, como a indução de fitoalexinas. As moléculas eliciadoras possuem a capacidade de induzir e ativar tais repostas, e assim sendo, técnicas têm buscado isolar e caracterizar frações com caráter eliciador. O trabalho teve por objetivo purificar, por cromatografia de troca iônica cromatografia de filtração em gel, moléculas eliciadoras a partir de nematoides fitopatogênicos, e testá-las na indução de faseolina em hipocótilos de feijoeiro e gliceolina em cotilédones de soja. O tampão Tris HCl 0,05 M (pH 6,8) foi utilizado como tratamento controle e o acibenzolar-S-metil (50 mg i.a. L-1) e o Saccharomyces cerevisiae (20 mg mL-1) foram utilizados como tratamento padrão de indução. Cromatografia de troca iônica (CTI) e cromatografia de filtração em gel (CFG) foram realizadas para separar frações com poder eliciador a partir de quinhentas fêmeas de nematoides de Meloidogyne incognita e Meloidogyne javanica. Para a purificação de eliciadores a patir de Meloidogyne javanica, por CTI, foram obtidos seis frações glicídicas e seis glicoproteicas. Estas, por sua vez, foram purificadas em CFG, sendo obtidos no total sessenta e três frações. As mesmas foram classificadas de acordo com sua natureza, sendo vinte e seis glicídicas e trinta e sete glicoproteicas, com massas moleculares variando de 29,19 a 2.989,25 kDa. Em relação a purificação de eliciadores de Meloidogyne incognita por CTI, foram obtidos nove frações glicídicas e cinco glicoproteicas. A partir destas, foram obtidos por CFG um total de cinquenta e oito frações, sendo vinte e cinco glicídicas e trinta e três glicoproteicas, com massas moleculares variando de 37,42 a 200,32 kDa. Das frações purificadas a partir de Meloidogyne javanica oito apresentaram potencial indutor de faseolina. Para gliceolina quinze frações mostraram efeito indutor. Em relação as frações purificadas a partir de Meloidogyne incognita, nenhuma fração apresentou potencial indutor de faseolina superior ao tratamento padrão. Entretanto, vinte e duas frações suprimiram a atividade de indução de fitoalexina. Para gliceolina dez frações induziram a mesma, enquanto que, vinte e três frações suprimiram a indução da gliceolina. A cromatografia foi eficiente na purificação de compostos eliciadores. Compostos com características supressoras de gliceolina e faseolina foram verificadas nos bioensaios. Para aquelas frações obtidas por CTI e posteriormente submetidas a CFG que não induziram fitoalexina, sugere-se que as moléculas necessitam atuar juntas para haver efeito eliciador e assim induzir a resposta de defesa no vegetal
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Baba, Hamed Mohamed Bey. "Purification et caractérisation de protéases alcalines des larves de Galleria Mellonella." Rouen, 1986. http://www.theses.fr/1986ROUES053.

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Deux protéases alcalines P1 et P2 ont été isolées à partir d'homogénats de larves de Galleria mellonella. Les 2 enzymes ont été séparées par chromatographie sur échangeurs d'ions et purifiées ultérieurement par filtration sur gel. Les 2 protéases se distinguent par leur pH optimum d'action, leur poids moléculaire et leur sensibilité vis-a-vis de différents inhibiteurs de protéases. La répartition anatomique de l'activité protéolytique montre la présence de 2 protéases alcalines similaires à P1 et P2 dans le tube digestif, et d'une protéase similaire à P1 dans le tissu adipeux
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Meinerz, Cristiane Claudia. "Indução de mecanismos bioquímicos de defesa em sorgo (Sorghum bicolor) por frações obtidas do decocto de avenca (Adiantum capillus-veneris)." Universidade Estadual do Oeste do Paraná, 2010. http://tede.unioeste.br:8080/tede/handle/tede/1413.

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Induction of resistance involves the activation of plant defense mechanisms in response to treatment with biotic or abiotic elicitors. The application of plant extracts in order to induce resistance mechanisms is an interesting alternative to chemical control, however, besides the presence of inducers, can occur the presence of suppressors. This study aimed to partially purificate through gel filtration chromatography (GFC) and precipitation with ammonium sulfate (SA), compounds present in decoct of Adiantum capillus-veneris, capable to induce defense mechanisms in sorghum mesocotyls, including phytoalexins and peroxidase, polyphenoloxidase (PPO), phenylalanine ammonia-lyase (PAL) and chitinase. The decoct 1% was fractionated with concentrations of ammonium sulfate, 0-20%, 20-40%, 40-60%, 60-80% and 80-100% of SA and those fractions were subjected to GFC. We obtained nine protein peaks and one glucosic peak for decoct with molecular weights ranging from 0.61 to 0.01 KDa; to fraction 0-20% were obtained two protein and two glucosic peaks, with molecular weights lower than 0.01 KDa, and concentration of sugars ranging from 4.1 to 17.5 mg mL-1; to fraction 20-40% were obtained three protein peaks (0.98 to 111.5 KDa) and five glucosic peaks (11.3 to 73.7 mg mL-1); to fraction 40-60% were obtained two protein peaks (0.09 to 111.5 KDa) and two glucosic peaks (5.6 to 7.5 mg mL-1); to fraction 60-80% were obtained six protein peaks (lower than 0.02 KDa) and two glucosic peaks (16.5 to 51.3 mg mL-1); and to fraction 80-100% were obtained three protein peaks (lower than 0.09 KDa). Sorghum mesocotyl were treated with fractions from the GFC, and decoct, acibenzolar-S-methyl (ASM) (125 mg L-1 of a. i. as elicitor of reference) and sodium phosphate buffer 10 mM pH 6.0. After incubation of 96 h were measured the levels of phytoalexins in mesocotyls and the activity of defense-related enzymes in leaves. Treatment with peak II (0,09 KDa) induced phytoalexin 6.68% more than. Among the fractionn, 60-80% increased 76% compared to ASM. To peroxidase the peak IV (lower than 0,01 KDa) increased 21% the activity compared to control water, and 44% compared to ASM. For the fraction 0-20% the protein peak II (lower than 0,01 KDa) increased 39% the activity in relation to the fraction 0-20% and 19% in relation to decoct. The fraction, 80-100% increased 89% compared to, ASM. For the PPO the peak VI (lower than 0,01 KDa) from decoct decreased 88% the activity compared to ASM. For PAL the peak II (lower than 0,01 KDa) from fraction 0-20% was 91% higher than decoct. For chitinase 1% peak IV (lower than 0,01 KDa) from decoct was 68% higher than the ASM. It was possible to induce defense mechanisms in sorghum by the application of partially purified fractions from A. capillus-veneris, which can allow to obtain new molecules and development alternative methods to control plant diseases
A indução de resistência envolve a ativação de mecanismos de defesa latentes existentes nas plantas em resposta ao tratamento com agentes bióticos ou abióticos. A aplicação de extratos vegetais visando à indução de mecanismos de resistência é uma alternativa interessante ao controle químico, entretanto, nestes extratos pode ocorrer além da presença de indutores, a presença de supressores. Este trabalho teve por objetivo a purificação parcial, por meio de cromatografia de filtração em gel (CFG) e precipitação com sulfato de amônio (SA), de compostos presentes em decocto de avenca (Adiantum capillus-veneris), eficientes na indução de mecanismos de defesa em mesocótilos de sorgo, incluindo as fitoalexina deoxiantocianidinas e as proteínas peroxidase, polifenoloxidase, fenilalanina amônia-liase e quitinase, buscando selecionar frações potencialmente eficientes na indução de resistência em plantas. Decocto (EA 1%) de A. capillus-veneris foi fracionado com concentrações de sulfato de amônio de 0-20%, 20-40%, 40-60%, 60-80% e 80-100% e esses cortes foram submetidos à cromatografia de filtração em gel (CFG). Foram obtidos nove picos protéicos e um pico glicídico para EA 1% com massas moleculares variando de 0,61 à 0,01 KDa; no corte 0-20% foram obtidos dois picos protéicos e dois glicídicos, com massas moleculares menores que 0,01 KDa, e concentração de açúcares redutores variando de 4,1 a 17,5 µg mL-1; no corte 20-40% três picos protéicos (111,5 à 0,98 KDa) e cinco glicídicos (11,3 a 73,7 µg mL-1 de açúcares); no corte 40-60% dois picos protéicos (111,5 à 0,09 KDa) e dois glicídicos (5,6 a 7,5 µg mL-1); no corte 60-80% seis picos protéicos (menor que 0,02 KDa) e dois glicídicos (16,5 a 51,3 µg mL-1); e no corte 80-100% três picos protéicos (menor que 0,09 KDa). Mesocótilos de sorgo foram tratados com as frações provenientes da CFG, além do decocto a 1%, acibenzolar-S-metil (ASM) (125 mg. L-1 do i.a. como elicitor de referência) e tampão fosfato de sódio 10 mM pH 6,0, totalizando 42 tratamentos. Após incubação por um período de 96 h, avaliou-se dos teores de fitoalexinas nos mesocótilos e análises bioquímicas dos folíolos. O tratamento pico II (0,09 KDa) do EA 1% mostrou-se eficiente na indução de fitoalexinas, sendo superior em 6,68% ao ASM. Entre os cortes, 60-80% permitiu incremento de 76% em relação ao ASM. Para peroxidase o pico IV (menor que 0,01 KDa) do EA 1% incrementou 21% a atividade em relação a testemunha água e 44% ao ASM. Para os precipitados 0-20% o pico protéico II (menor que 0,01 KDa) promoveu incremento de 39% na atividade em relação ao corte 0-20% e 19% para o EA 1%. O precipitado 80-100% foi superior 89% ao ASM. Para polifenoloxidase o pico protéico VI (menor que 0,01 KDa) do EA1% reduziu 88% a atividade em relação ao ASM. Para fenilalanina amônia-liase o pico protéico II (menor que 0,01 KDa) do corte 0-20% foi 91% superior ao EA 1%. Para quitinase o pico protéico IV (menor que 0,01 KDa) do EA 1% foi 68% superior ao ASM. Foi possível induzir mecanismos de defesa em sorgo pela aplicação de frações parcialmente purificadas de A. capillus-veneris, o que pode permitir a obtenção de novas moléculas e o desenvolvimento de métodos alternativos para controle de doenças em plantas
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Kruger, Sarah Jane, and n/a. "Characterisation of Proteins from Grevillea robusta and NMR Studies of the Serine Protease Inhibitor." Griffith University. School of Science, 2004. http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20040618.150708.

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Proteins that recognise the sugar surface structures on cells have an enormous potential to be used as tools in the characterisation of these structures. A group of proteins, called lectins, have been identified that can bind to carbohydrate complexes on the receptors of cells. The crude extract from Grevillea robusta seeds was found to contain lectin-like proteins that were different from most other lectins, as they would specifically target the receptors of white blood cells and not those found on red blood cells. Therefore, the lectin isolated from G.robusta could be used as a tool to identify the specific surface structures on white blood cells. The lectin was isolated using affinity chromatography where a complex (oligosaccharide) matrix was used. Agglutination, binding and sugar inhibition assays confirmed the isolated protein was a lectin. The lectin was found in low amounts (up to 5% of the total protein content) within the seeds of G.robusta. As a result of this low yield, the identification of the lectin by PAGE was difficult because the levels of protein were beyond the detection limit of the commercial staining reagents. The lectin was called the GR2 protein and was characterised as a monocot mannose binding lectin based on its sugar specificity for only mannose. A serine protease inhibitor was isolated from the seeds of G.robusta using two different chromatography methods, reverse phase HPLC (GR1.HPLC) and gel filtration chromatography (GR1.GF). Ion exchange chromatography was used to initially separate the proteins in the crude extract and the fraction containing the GR1 protein was further purified using reverse phase HPLC (GR1.HPLC). N-terminal sequencing results of the GR1.HPLC protein, showed evidence of proteolytic cleavage during the extraction process, which lead to the second purification method being established. Protease inhibitors were added to the buffers prior to being purified by gel filtration chromatography, which resulted in the GR1 protein being isolated from the crude extract without the presence of the contaminating protein. Mass spectroscopy identified the molecular weight of the GR1 protein to be 6669Da and the full amino acid sequence was derived by cDNA techniques. Sequence alignment studies of the GR1 protein showed significant similarities with the Bowman-Birk inhibitor. The positioning of the cysteine residues were conserved throughout the Bowman-Birk superfamily, however these residues were not conserved within the GR1 protein. Competitive inhibition assays on the GR1 protein revealed the protein could inhibit both trypsin and chymotrypsin at similar levels to that seen for the Bowman-Birk inhibitor. Therefore, the GR1 protein was characterised as a member of the Bowman-Birk superfamily of serine protease inhibitors. The three-dimensional structure of the GR1 protein was determined using two-dimensional NMR spectroscopy. Computer programs such as XEASY, DYANA and SYBYL® were used to tabulate the information taken from the 2D experiments, generate structures and minimise these structures respectively. The solution structure of the GR1 protein was found to contain a region of antiparallel β-sheet structure that corresponded to the trypsin binding site and the remainder of the protein consisted of loops and turns that were held together by disulfide bridges (the chymotrypsin-binding region). Structural similarities between the GR1 protein and the Bowman-Birk inhibitor existed only in the trypsin-binding site of the Bowman-Birk inhibitor. The GR1 protein is the first member of the Proteaceae family to be characterised as a Bowman-Birk inhibitor. This thesis outlines the isolation and biochemical characterisation of the two proteins found within Grevillea robusta and also describes the steps involved and results obtained in determining the three-dimensional structure of the GR1 protein.
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Kruger, Sarah Jane. "Characterisation of Proteins from Grevillea robusta and NMR Studies of the Serine Protease Inhibitor." Thesis, Griffith University, 2004. http://hdl.handle.net/10072/366534.

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Proteins that recognise the sugar surface structures on cells have an enormous potential to be used as tools in the characterisation of these structures. A group of proteins, called lectins, have been identified that can bind to carbohydrate complexes on the receptors of cells. The crude extract from Grevillea robusta seeds was found to contain lectin-like proteins that were different from most other lectins, as they would specifically target the receptors of white blood cells and not those found on red blood cells. Therefore, the lectin isolated from G.robusta could be used as a tool to identify the specific surface structures on white blood cells. The lectin was isolated using affinity chromatography where a complex (oligosaccharide) matrix was used. Agglutination, binding and sugar inhibition assays confirmed the isolated protein was a lectin. The lectin was found in low amounts (up to 5% of the total protein content) within the seeds of G.robusta. As a result of this low yield, the identification of the lectin by PAGE was difficult because the levels of protein were beyond the detection limit of the commercial staining reagents. The lectin was called the GR2 protein and was characterised as a monocot mannose binding lectin based on its sugar specificity for only mannose. A serine protease inhibitor was isolated from the seeds of G.robusta using two different chromatography methods, reverse phase HPLC (GR1.HPLC) and gel filtration chromatography (GR1.GF). Ion exchange chromatography was used to initially separate the proteins in the crude extract and the fraction containing the GR1 protein was further purified using reverse phase HPLC (GR1.HPLC). N-terminal sequencing results of the GR1.HPLC protein, showed evidence of proteolytic cleavage during the extraction process, which lead to the second purification method being established. Protease inhibitors were added to the buffers prior to being purified by gel filtration chromatography, which resulted in the GR1 protein being isolated from the crude extract without the presence of the contaminating protein. Mass spectroscopy identified the molecular weight of the GR1 protein to be 6669Da and the full amino acid sequence was derived by cDNA techniques. Sequence alignment studies of the GR1 protein showed significant similarities with the Bowman-Birk inhibitor. The positioning of the cysteine residues were conserved throughout the Bowman-Birk superfamily, however these residues were not conserved within the GR1 protein. Competitive inhibition assays on the GR1 protein revealed the protein could inhibit both trypsin and chymotrypsin at similar levels to that seen for the Bowman-Birk inhibitor. Therefore, the GR1 protein was characterised as a member of the Bowman-Birk superfamily of serine protease inhibitors. The three-dimensional structure of the GR1 protein was determined using two-dimensional NMR spectroscopy. Computer programs such as XEASY, DYANA and SYBYL® were used to tabulate the information taken from the 2D experiments, generate structures and minimise these structures respectively. The solution structure of the GR1 protein was found to contain a region of antiparallel β-sheet structure that corresponded to the trypsin binding site and the remainder of the protein consisted of loops and turns that were held together by disulfide bridges (the chymotrypsin-binding region). Structural similarities between the GR1 protein and the Bowman-Birk inhibitor existed only in the trypsin-binding site of the Bowman-Birk inhibitor. The GR1 protein is the first member of the Proteaceae family to be characterised as a Bowman-Birk inhibitor. This thesis outlines the isolation and biochemical characterisation of the two proteins found within Grevillea robusta and also describes the steps involved and results obtained in determining the three-dimensional structure of the GR1 protein.
Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Science
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Books on the topic "Gel filtration chromatography"

1

Harkin, J. M., Helmut Determann, and E. Gross. Gel Chromatography Gel Filtration · Gel Permeation · Molecular Sieves: A Laboratory Handbook. Springer London, Limited, 2012.

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Determann, Helmut. Gel Chromatography : Gel Filtration · Gel Permeation · Molecular Sieves: A Laboratory Handbook. Springer London, Limited, 2013.

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Determann, Helmut. Gel Chromatography Gel Filtration · Gel Permeation · Molecular Sieves: A Laboratory Handbook. Springer, 2012.

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Determann, Helmut. Gel Chromatography: Gel Filtration · Gel Permeation · Molecular Sieves a Laboratory Handbook. Springer London, Limited, 2012.

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Kirkland, Joseph J., Andre Striegel, Wallace W. Yau, and Donald D. Bly. Modern Size-Exclusion Liquid Chromatography: Practice of Gel Permeation and Gel Filtration Chromatography. Wiley & Sons, Incorporated, John, 2009.

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Kirkland, Joseph J., Andre Striegel, Wallace W. Yau, and Donald D. Bly. Modern Size-Exclusion Liquid Chromatography: Practice of Gel Permeation and Gel Filtration Chromatography. Wiley & Sons, Limited, John, 2009.

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Book chapters on the topic "Gel filtration chromatography"

1

Ó’Fágáin, Ciarán, Philip M. Cummins, and Brendan F. O’Connor. "Gel-Filtration Chromatography." In Methods in Molecular Biology, 25–33. Totowa, NJ: Humana Press, 2010. http://dx.doi.org/10.1007/978-1-60761-913-0_2.

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Ó’Fágáin, Ciarán, Philip M. Cummins, and Brendan F. O’Connor. "Gel-Filtration Chromatography." In Methods in Molecular Biology, 15–25. New York, NY: Springer New York, 2016. http://dx.doi.org/10.1007/978-1-4939-6412-3_2.

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Moore, Peter M. "Gel Filtration Chromatography." In Adsorption: Science and Technology, 561–76. Dordrecht: Springer Netherlands, 1989. http://dx.doi.org/10.1007/978-94-009-2263-1_29.

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Kummari, Raghupathi, and Kakoli Bose. "Gel Filtration Chromatography." In Textbook on Cloning, Expression and Purification of Recombinant Proteins, 199–219. Singapore: Springer Singapore, 2022. http://dx.doi.org/10.1007/978-981-16-4987-5_8.

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Hagel, Lars. "Gel Filtration: Size Exclusion Chromatography." In Methods of Biochemical Analysis, 51–91. Hoboken, NJ, USA: John Wiley & Sons, Inc., 2011. http://dx.doi.org/10.1002/9780470939932.ch3.

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Page, Mark, and Robin Thorpe. "Purification of IgG Using Gel-Filtration Chromatography." In Springer Protocols Handbooks, 735–37. Totowa, NJ: Humana Press, 1996. http://dx.doi.org/10.1007/978-1-60327-259-9_131.

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Booy, Evan P., Hui Meng, and Sean A. McKenna. "Native RNA Purification by Gel Filtration Chromatography." In Recombinant and In Vitro RNA Synthesis, 69–81. Totowa, NJ: Humana Press, 2012. http://dx.doi.org/10.1007/978-1-62703-113-4_6.

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Bai, Yan. "Detecting Protein-Protein Interactions by Gel Filtration Chromatography." In Methods in Molecular Biology, 223–32. New York, NY: Springer New York, 2015. http://dx.doi.org/10.1007/978-1-4939-2425-7_13.

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Elwell, Lynn P. "A Rapid Method for Purifying Escherichia coli β-galactosidase Using Gel-Filtration Chromatography." In Filtration and Purification in the Biopharmaceutical Industry, 467–80. Third edition. | Boca Raton, Florida : CRC Press, 2019. | Series: Drugs and the pharmaceutical sciences: CRC Press, 2019. http://dx.doi.org/10.1201/9781315164953-18.

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Masoodi, Khalid Z., Sameena Maqbool Lone, and Rovidha Saba Rasool. "Gel-filtration or size-exclusion chromatography." In Advanced Methods in Molecular Biology and Biotechnology, 147–49. Elsevier, 2021. http://dx.doi.org/10.1016/b978-0-12-824449-4.00026-8.

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Conference papers on the topic "Gel filtration chromatography"

1

Xiong, Yi-min, Zhen-yi Wang, Ye-lu Xu, and Chenq-wu Chi. "REVERSIBLE BINDING OF THE TISSUE PLASMINOGEN ACTIVATOR WITH FIBRONECTIN." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644404.

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Fresh pig heart tissues were homogenized and deli-pidized with cold acetone. The dry acetone powder was extracted with 0.45 M potassium acetate pH 4.5, the extract was fractionated with ammonium sulfate to 60% saturation, successively followed by four chromatography steps: chromatography on CM-Sepharose CL-6B; gel filtration on Sephadex G-100; Fibrin-Sepharose affinity chromatography and chromatography on DEAE-Sepharose CL-6B. The fibrin plate method was used for the tissue plasminogen activator (t-PA) activity determination. The porcine t-PA purified was proved to be homogeneous either by SDS gel electrophoresis or by high performance liguid chromatography with a molecular weight of 67,000. Using the human melanoma t-PA antigen and its rabbit antiserum, the double immunodiffusion and immu-noelectrophoretic tests demonstrated that the porcine t-PA possessed a cross-reaction with the human t-PA. During the purification procedure, it was found that the porcine t-PA was contaminated and reversibly bound with the cell fibronectin. The binding ability depends on salt concentration. They could be separated from each other on Sepharose G-100 gel filtration at high salt concentration, this reversible binding was further confirmed by mixing these two purified components monitored on high performance liquid chromatography. The fibronectin-bound t-PA retained its ability to activate plasminogen, indicating that these two proteins may exist in a form of complex in the in vivo tissue. Some monoclonal antibodies against the porcine t-PA were obtained, being used to screen what domain of the enzyme is responsible for binding the cell fibronectin.
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Carlsson, I., J. Chmielewska, and B. Wiman. "ON DIFFERENT MOLECULAR EORMS OE PLASMINOGEN ACTIVATOR INHIBITOR." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644434.

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The production of plasminogen activator inhibitor (PAI) by the human cell-lines Hep G2 and HT 1080 have been studied by immunochemical and functional methods. In conditioned medium collected after 2h, the PAI seemed to be almost fully active, but with increasing incubation time the activity was gradually lost, in spite of that the PAI-antigen content increased continously. The active PAI form can be separated from the inactive form by gel-filtration. The inactive form behaves as a low Mr (about 50,000) component in the absence and in the presence of sodium dodecyl-sulphate. In contrast, the active form of PAI behaves as a high Mr (>300,000) compound in the absence of sodium dodecylsulphate but as a low MT compound in its presence. The low M_r inactive PAI has been purified to homogeneity from HT 1080 conditioned medium, collected in the absence of fetal calf serum. This was achieved by chromatography on Concanavalin A-Sepharose, gel-filtration on Sephacryl S-300 and affinity chromatography on insolubilized monoclonal antibodies against PA-inhibitor. On treatment of this form of the inhibitor with 4 mol/L Guanidinium chloride, the activity was regained, but its gel-filtration behaviour was unchanged in the absence of serum/plasma (Mr about 50,000). Addition of plasma or serum prior to the gel-filtration, changed the elution pattern of PAI towards a high Mr form. The reason for this behaviour is not yet fully understood, but the most plausible explanation is the presence of a high Mr PAI-binding protein in plasma/serum. This hypothesis is presently being explored .
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Stachowiak, Jeanne C., Erin E. Shugard, Pamela Caton, Bruce P. Mosier, Ron Renzi, Rafael V. Davalos, Gregory J. McGraw, Blake A. Simmons, Victoria A. Vandernoot, and Brent A. Haroldsen. "Automated Sample Preparation System for Rapid Biological Threat Detection." In ASME 2005 International Mechanical Engineering Congress and Exposition. ASMEDC, 2005. http://dx.doi.org/10.1115/imece2005-80945.

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Rapid, automated sample preparation of bacterial cells and spores is required for threat analysis by remotely deployed chemical and biological warning systems. Sandia is designing, building, and testing an automated front-end sample preparation system based on miniature and microfluidic components, with the goal of concentrating bacterial species collected from the air, harvesting and solubilizing proteins from them, and delivering them to Sandia’s MicroChemLab capillary gel electrophoresis system1,2 for analysis (Fig. 1). Miniature, motorized valves and pumps control flow between system components connected by fused silica capillaries (Fig. 4). Sample processing modules include concentration by dielectrophoresis in an array of insulating posts or by mechanical filtration; heat-activated chemical lysis; mechanical filtration; removal of chemical lysis agents by size exclusion chromatography (SEC); and in-capillary fluorescent labeling.
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Wiman, B., T. Carlsson, and J. Chmielewska. "EVIDENCE FOR A PLASMINOGEN ACTIVATOR INHIBITOR BINDING PROTEIN IN PLASMA." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642859.

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For several years it has been known that plasminogen activator inhibitor in plasma behaves as a high molecular weight compound on gelfiltration, in spite of that the molecular weight is only 50,000 in the presence of sodium dodecylsul-phate. The reason for this has so far been unknown. On gelfiltration of plasma, to which purified latent PAI from HT 1080 cells was added, the PAI antigen gel-filtered as a 50,000 Mr protein. However, if the latent form of PAI was reactivated by guanidinium chloride prior to the gel-filtra-tion experiment, an apparent molecular weight of about 250.000 for PAI antigen and activity was observed. If more than 10,000 U of PAI activity was added/mL of normal human plasma, excess PAI occurred at 50,000 Mr on gel-filtration. Human normalplasma was subjected to gel-filtration on sepha-cryl S-300 or Sepharose 6B and the fractions were checked for capacity to transform low Mr functional PAI to high Mr functional PAI. This capacity was only found in the 150 - 200,000 Mr region of the chromatogram. These data suggest that human plasma contains a protein that binds active forms of PAI. The complex of this protein and PAI could be dissociated by gelfiltration in the presence of 3 mol/L guanidinium chloride or 0.1 % (w/v) sodium dodecylsulphate. The physiological or pathophysiological role of the PAI-binding protein is not known. Work with purification of the protein is in progress. Considerable purification have so far been obtained by precipitation with polyethylenglycol 6000 (0-6%), gel-filtration on Sephacry1 S-300, followed by affinity chromatography on heparin-Sepharose.
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Nurhidayat, I., S. Setiasih, S. Handayani, and S. Hudiyono. "Kinetic studies of bromelain purified from Palembang pineapple (Ananas comosus [L.] Merr) using gel filtration chromatography and its activity as antiplatelet aggregation." In PROCEEDINGS OF THE 3RD INTERNATIONAL SYMPOSIUM ON CURRENT PROGRESS IN MATHEMATICS AND SCIENCES 2017 (ISCPMS2017). Author(s), 2018. http://dx.doi.org/10.1063/1.5064065.

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6

Morinaga, T., Y. Itagaki, A. Suzuki, H. Yasuda, and K. Higashio. "PURIFICATION AND CHARACTERIZATION OF TISSUE PLASMINOGEN ACTIVATOR PRODUCED BY IMR-90 CELLS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644393.

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Tissue plasminogen activator ( t-PA ) produced by IMR-90 ( human embryonic fibroblast ) cells cultured in the serum-free medium ( DMEM ) containing 1% proteos^e peptone and 1.6 - 3.6mM CaCl2 was purified by the procedure consisted of ultrafiltration, immunoadsorpt ion chromatography, HPLC and lysine-Sepharose chromatography. The yield of t-PA from the culture broth was approximately 47%. The purified t-PA migrated as a single band on SDS-polyacrylamide gels. The molecular weight of the t-PA was estimated to be 66,000 by SDS-polyacrylamide gel electrophoresis and 69,000 by gel filtration method. Purified t-PA had a specific activity of 36 × 104 IU/mg protein by fiblin plate method or 54 - 56 X 104 IU/mg protein by clot lysis method using t-PA obtained from WHO as a standard. The amino acid composition of fibroblast t-PA was very similar to those of melanoma t-PA and uterine t-PA. Isoelectric point of fibroblast t-PA ranged from 5-7 to 8.2. The t-PA had twice as much affinity for fiblin as did high molecular weight urokinase ( UK ). Both t-PA and UK had optimum temperature at 41°C and optimum pH between 8.0 - 9.0. The polyclonal and monoclonal antibodies raised against t-PA quenched t-PA activity but had no effect on UK activity. The inhibitors of serine proteases, difluorophos-phate and gabexate mesilate, strongly inhibited the activities of fibroblast t-PA and UK. The nucleotide sequence analysis of the t-PA cDNA isolated from the cDNA library prepared from IMR-90 mRNA revealed the nucleotide changes at two positions in the coding region as compared to that of melanoma t-PA cDNA. Neither of the changes replaced the coded amino acid. The N-terminal amino acid of fibroblast t-PA was determined to be valine, indicatig the structural similarity of fibroblast t-PA to uterine t-PA.
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7

Masci, P. P., A. N. Whitaker, J. J. Morrison, and E. A. Bennett. "PURIFICATION AND CHARACTERIZATION OF THE PROCOAGULANT OF THE VENOM OF TROPIDECHIS CARINATUS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644322.

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Tropidechis carinatus is a venomous elapid snake distributed throughout Eastern Queensland. It has been considered as a tropical relative of Notechis scutatus and, similarly, the crude venom contains an indirect prothrombin activator, which will clot plasma provided that Factor V is present. Myotoxins and neurotoxins are also present. Envenomated patients regularly develop disseminated intravascular coagulation. The crude whole venom of T.carinatus was shown to have five major components by gel filtration, SDS PAGE and HPLC, and even more components by isoelectric focusing. The procoagulant eluted with a molecular weight of 55,000, being found in peak II on gel filtration on Sephadex-G150. The procoagulant was purified using a combination of Sephadex-G150 chromatography and ion-exchange on DEAE Sephadex-A50 and shown to migrate as a single band of molecular weight 55.000 by SDS PAGE. On reduction by β -mercaptoethanol this component was resolvec into u heavy chain of molecular weight 30.000 and a light chain of 25,000. The procoagulant was shown to bind to con A-Sepharose 4B and Blue Sepharose 4B. Coagulation studies using this purified procoagulant confirmed a factor Xa-like activity activating prothrombin in the presence of factor V. The purified fraction is unstable in buffer solutions at 4°C, probably because of trypsin - like autodigestion. Ouchterlony studies of the procoagulants of T.carinatus and N.scutatus show both lines of homogeneity and spurring, indicating similarities but also significant differences between the two proteins. The purified procoagulant was lethal to adult rats, an IV injection of 10 μg killing in 1 - 2 minutes. Death was prevented by prior heparinization, suggesting that the procoagulant is toxic in the absence of neurotoxin and other components.
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Gargan, P. E., and V. A. Ploplis. "IDENTIFICATION AND PURIFICATION OF AN INHIBITOR TO PLASMINOGEN ACTIVATORS FROM PROSTATE ADENOCARCINOMA CELLS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643192.

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The rat prostate adencarcinoma cell line (PA-III), derived from qerm-free Lobund Wistar rats has served as a unique model for prostatic cancer. The supernatant from confluent cultures of the cell line was shown by zymographic analysis to contain three molecular forms of plasminogen activator. One of the activators was characterized in a previous study and has been classified as a tissue type plasminogen activator(Mr=30,000). In the same study the activator with a Mr=45,000 was shown to be of the urokinase class. The high molecular weight(Mr=120, 000) activator, which has not been described previously, was identified as a complex between the tissue plasminogen activator and an inhibitor. Dissociation of the activator and its inhibitor was achieved by incubation of the complex with ammonium hydroxide (1.5M) in the presence of NaDodGO4 (2%). Treatment with NaDodSO4 alone did not dissociate the complex but did however enhance its fibrinolytic activity, suggesting disruption of hydropnotic interactions leading to exposure of the activators active site.A scheme for the purification of the two plasminogen activators, the activator-inhibitor complex, and the free inhibitor was developed. The various components were purified approximately 1000 fold by hydrophobic interaction chromatography on Phenyl-Sepharose, affinity chromatography on Con A Sepharose and by gel filtration on Sephacryl S-200. The purified inhibitor was capable of inhibiting the fibrinolytic activity of both the urokinase type and tissue type plasminogen activators produced by the PAIII cells.
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Gheysen, D., L. Piérard, P. Jacobs, H. R. Lijnen, A. Bollen, and D. Collen. "PROPERTIES OF A HUMAN RECOMBINANT FUSION PROTEIN OF THE ‘FINGER’ DOMAIN OF TISSUE-TYPE PLASMINOGEN ACTIVATOR (t-PA) AND A TRUNCATED SINGLE CHAIN UROKINASE-TYPE PLASMINOGEN ACTIVATOR (scu-PA)." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643941.

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A hybrid between human tissue-type plasminogen activator (t-PA) and human single chain urokinase-type plasminogen activator (scu-PA) was obtained by ligation of cDNA fragments encoding the NH2-terminal amino acids 1 to 67 of t-PA and the COOH-ter-minal amino acids 136 to 411, of scu-PA. Both this chimaeric cDNA and cDNA encoding scu-PA were expressed in a mammalian system (HAK-cells) using bovine papilloma virus (BPV) derived vectors. Two stable cell lines were obtained which secreted the recombinant hybrid and the scu-PA at 1 μg/ml and 2 μg/ml u-PA related antigen respectively into the culture medium. Following purification by Zinc chelate Sepharose, immunoadsorption chromatography, benzamidine-Sepharose and Ultrogel AcA44 gel filtration, highly purified proteins were obtained with a yield of about 200 μg/1. SDS gel electrophoresis under reducing conditions showed single bands with M 43,000 and M 50,000 respectively. Following conversion to urokinase with plasmin, both proteins had a specific amidolytic activity comparable to that of natural scu-PA. Both proteins activated plasminogen directly with km1.4 and 0.5 μM and k2 0.0034 s and 0.0027 s . Neither protein bound specifically to fibrin.Thus the fusion of the finger-like domain of t-PA to the COOH-terminal part of scu-PA does not confer fibrin affinity of t-PA to this chimaeric protein. However, peptide material can be fused to the COOH-terminal part of scu-PA without perturbing its enzymatic properties.
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Müller, E., A. Henschen, and G. Wunderer. "IDENTIFICATION OF A NEW HUMAN KININ, ILE-SER-BRADYKININ, BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY AND SEQUENCE ANALYSIS IN OVARIAN CARCINOMA ASCITES." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642848.

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Human blood has been shown to contain two different kinin precursors, i.e0 the high and the low molecular mass kininogen0 These two kininogens release the same kinins, with the starting sequences Met-Lys-Arg-Pro-, Lys-Arg-Pro- or just Arg-Pro-depending on the releasing enzyme. The kinin starting with Arg-Pro- is denoted as bradykinin. In rats a different kininogen, called T-kininogen, is also present, especially as the major acute-phase protein in this species. The corresponding kinin, T-kinin, has the starting sequence Ile-Ser-Arg-Pro-. This type of kininogen or kinin has previously never been detected in human tissues. However, during the course of the present study evidence for existance of a third human kininogen, giving rise to human T-kinin, was obtained.Ascites from patients with metastatic ovarian carcinoma has been shown to contain high amounts of vascular permeability-increasing activity as determined by a rat skin-Evans blue test. When the ascites was fractionated by gel filtration followed by reversed-phase high-performance liquid chromatography (HPLC) a component could be isolated which by its total sequence and amino acid composition was identified as Ile-Ser-bradykinin. Several degradation products of this kinin were also detected as separate components in the chromatographies. The human Ile-Ser-bradykinin appeared on reversed-phase HPLC in the same position as synthetic T-kinin. It could be differentiated in this chromatography system from Met-Lys-bradykinin, Lys-bradykinin and bradykinin. It may be assumed that human Ile-Ser-bradykinin is released_ from a third, so far unidentified human kininogen which is only or predominantly expressed under certain pathophysiological conditions, and that therefore this new kinin might be employed as a tumor marker.
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