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Teizo Fujita - One of the best experts on this subject based on the ideXlab platform.

Daniela Diogenes De Carvalho - One of the best experts on this subject based on the ideXlab platform.

  • Caracterização molecular e atividade citotoxica sobre celulas tumorais da lectina do veneno da serpente Bothrops jararacussu
    [s.n.], 2018
    Co-Authors: Daniela Diogenes De Carvalho
    Abstract:

    Orientador : Jose Camillo NovelloTese (doutorado) - Universidade Estadual de Campinas, Instituto de BiologiaResumo: As lectinas são proteínas ligantes de carboidratos, de origem não-imune, encontradas numa grande variedade de organismos. Aquelas isoladas de venenos de serpentes são constituídas de cadeias polipeptídicas relativas à região molecular correspondente ao domínio de reconhecimento de carboidrato (CRD) de lectinas. Devido às interações lectina ¿ carboidrato serem reversíveis, e não covalentes, estas moléculas podem ser utilizadas como importantes ferramentas bioquímicas. Este estudo tem como objetivo: (i) a caracterização da estrutura primária da lectina do veneno da serpente Bothrops jararacussu (BJcuL); (ii) a investigação do efeito de BJcuL sobre a adesão de células tumorais a proteínas da matriz extracelular, (iii) e sobre a proliferação destas e de células endoteliais. O seqüenciamento N-terminal de BJcuL revelou a presença de uma única seqüência, indicando que esta proteína é composta de cadeias idênticas. A determinação da estrutura primária da lectina realizada por meio de análise dos peptídeos obtidos pela fragmentação da proteína com c1ostripaína e protease SV-8, demonstrou que BJcuL possui os 18 resíduos invariantes que caracterizam o tipo C de lectinas animais. Este fato indica que BJcuL pertence à família das lectinas tipo-C ligantes de I3-galactosídeos e apresenta símilaridades estruturais com a região C-terminal do CRD das lectinas animais de membrana. Nos primeiros testes em cultura de células tumorais, a viabilidade celular foi avaliada após 5 dias de cultivo na presença da lectina. Nestas condições, a BJcuL inibiu 50% do crescimento das células de carcinoma de rins ou pâncreas em concentrações entre 1 e 2 mM. Em testes posteriores, foi observado que as células de carcinoma de mama ou de ovário foram capazes de aderir fracamente a BJcuL. Entretanto, esta adesão não inibiu a fixação destas células à proteínas da matriz extracelular tais como; fibronectina, laminina e colágeno tipo I. Após proliferação de linhagens tumorais por 4 dias na presença de BJcuL, foi observado um efeito inibidor dose-dependente da lectina em células tumorais de glioma e carcinomas de mama ou ovário. No intuito de verificar as propriedades anti angiogênicas da lectina, foi realizado um ensaio de proliferação de células endoteliais na presença de BJcuL. Os resultados mostraram que a BJcuL, numa concentração de 0,09mM, foi capaz de reduzir em 50% a viabilidade destas células. Os resultados aqui mostrados fornecem base para estudos acerca da estrutura molecular da BJcuL e suas funções, da caracterização da ligação lectina-carboidrato, e dos mecanismos envolvidos na ligação da lectina às superficies celularesAbstract: Lectins are carbohydrate-binding proteins of non-immune origin found in a diverse array of organisms. Snake venom Lectins are malleable molecules, and their molecular structure comprises the carbohydrate recognition domain (CRD) characterized in other Ca2+-dependent animal Lectins. They could be used as interesting tools since lectin-glycan interactions are reversible and non-covalent. The aim of this study was (i) to characterize the molecular structure of lectin from the venom of the snake Bothrops jararacussu (BJcuL); (ii) to investigate the effect of BJcuL on the adhesion properties of tumor cells to the extracellular matrix proteins, (iii) and on the proliferation of cancer and endothelial cells.The determination of the single N-terminal sequence has shown that BJcuL is a homodimer. The analysis of the complete amino acid sequence of the peptides obtained by enzymatic BJcuL digestion (clostripain and SV-8 protease) showed that this lectin displays 18 invariant amino acid residues characteristics of C-type Lectins. This fact implies that BJcuL possesses structural similarities to the C-terminal region of the animal membrane Lectins CRD, belonging to the C-type j3-galactoside binding lectin family. In the first evaluation in a tumor cell system, cell viability was evaluated after 5 days cultivation. This lectin was a potent inhibitor of growth in human renal or carcinoma cells, with 50% inhibitory concentrations (lC50) of cell growth between 1 and 2 mM of BJcuL. In the second evaluation, cells of human metastatic breast cancer or human ovarian carcinoma cell lines weakly adhere to BJcuL. However, BJcuL was not capable of inhibiting adhesion of these cells to the extracellular matrix proteins such as fibronectin, laminin and type I collagen. 1t was also observed a dose-dependent inhibitory effect of BJcuL in proliferation assays with glioma, breast or ovarian carcinoma cells after 4 days of incubation. Proliferation assays were performed on bovine brain and endothelial cells in order to verify the antiangiogenic properties of the BJcuL These experiments revealed that BJcuL was capable to reduce cell proliferation (lC5o of 0,09mM). Taken together, our findings could be helpful to further studies regarding the lectin structure and function, lectin-glycan binding characterization, as well as, the mechanisms involved in BJcuL binding to cell surface molecules, which influence cell proliferation and angiogenesisDoutoradoBioquimicaDoutor em Biologia Funcional e Molecula

  • Caracterização molecular e atividade citotoxica sobre celulas tumorais da lectina do veneno da serpente Bothrops jararacussu
    Universidade Estadual de Campinas. Instituto de Biologia, 2002
    Co-Authors: Daniela Diogenes De Carvalho
    Abstract:

    As lectinas são proteínas ligantes de carboidratos, de origem não-imune, encontradas numa grande variedade de organismos. Aquelas isoladas de venenos de serpentes são constituídas de cadeias polipeptídicas relativas à região molecular correspondente ao domínio de reconhecimento de carboidrato (CRD) de lectinas. Devido às interações lectina ? carboidrato serem reversíveis, e não covalentes, estas moléculas podem ser utilizadas como importantes ferramentas bioquímicas. Este estudo tem como objetivo: (i) a caracterização da estrutura primária da lectina do veneno da serpente Bothrops jararacussu (BJcuL); (ii) a investigação do efeito de BJcuL sobre a adesão de células tumorais a proteínas da matriz extracelular, (iii) e sobre a proliferação destas e de células endoteliais. O seqüenciamento N-terminal de BJcuL revelou a presença de uma única seqüência, indicando que esta proteína é composta de cadeias idênticas. A determinação da estrutura primária da lectina realizada por meio de análise dos peptídeos obtidos pela fragmentação da proteína com c1ostripaína e protease SV-8, demonstrou que BJcuL possui os 18 resíduos invariantes que caracterizam o tipo C de lectinas animais. Este fato indica que BJcuL pertence à família das lectinas tipo-C ligantes de I3-galactosídeos e apresenta símilaridades estruturais com a região C-terminal do CRD das lectinas animais de membrana. Nos primeiros testes em cultura de células tumorais, a viabilidade celular foi avaliada após 5 dias de cultivo na presença da lectina. Nestas condições, a BJcuL inibiu 50% do crescimento das células de carcinoma de rins ou pâncreas em concentrações entre 1 e 2 mM. Em testes posteriores, foi observado que as células de carcinoma de mama ou de ovário foram capazes de aderir fracamente a BJcuL. Entretanto, esta adesão não inibiu a fixação destas células à proteínas da matriz extracelular tais como; fibronectina, laminina e colágeno tipo I. Após proliferação de linhagens tumorais por 4 dias na presença de BJcuL, foi observado um efeito inibidor dose-dependente da lectina em células tumorais de glioma e carcinomas de mama ou ovário. No intuito de verificar as propriedades anti angiogênicas da lectina, foi realizado um ensaio de proliferação de células endoteliais na presença de BJcuL. Os resultados mostraram que a BJcuL, numa concentração de 0,09mM, foi capaz de reduzir em 50% a viabilidade destas células. Os resultados aqui mostrados fornecem base para estudos acerca da estrutura molecular da BJcuL e suas funções, da caracterização da ligação lectina-carboidrato, e dos mecanismos envolvidos na ligação da lectina às superficies celularesLectins are carbohydrate-binding proteins of non-immune origin found in a diverse array of organisms. Snake venom Lectins are malleable molecules, and their molecular structure comprises the carbohydrate recognition domain (CRD) characterized in other Ca2+-dependent animal Lectins. They could be used as interesting tools since lectin-glycan interactions are reversible and non-covalent. The aim of this study was (i) to characterize the molecular structure of lectin from the venom of the snake Bothrops jararacussu (BJcuL); (ii) to investigate the effect of BJcuL on the adhesion properties of tumor cells to the extracellular matrix proteins, (iii) and on the proliferation of cancer and endothelial cells.The determination of the single N-terminal sequence has shown that BJcuL is a homodimer. The analysis of the complete amino acid sequence of the peptides obtained by enzymatic BJcuL digestion (clostripain and SV-8 protease) showed that this lectin displays 18 invariant amino acid residues characteristics of C-type Lectins. This fact implies that BJcuL possesses structural similarities to the C-terminal region of the animal membrane Lectins CRD, belonging to the C-type j3-galactoside binding lectin family. In the first evaluation in a tumor cell system, cell viability was evaluated after 5 days cultivation. This lectin was a potent inhibitor of growth in human renal or carcinoma cells, with 50% inhibitory concentrations (lC50) of cell growth between 1 and 2 mM of BJcuL. In the second evaluation, cells of human metastatic breast cancer or human ovarian carcinoma cell lines weakly adhere to BJcuL. However, BJcuL was not capable of inhibiting adhesion of these cells to the extracellular matrix proteins such as fibronectin, laminin and type I collagen. 1t was also observed a dose-dependent inhibitory effect of BJcuL in proliferation assays with glioma, breast or ovarian carcinoma cells after 4 days of incubation. Proliferation assays were performed on bovine brain and endothelial cells in order to verify the antiangiogenic properties of the BJcuL These experiments revealed that BJcuL was capable to reduce cell proliferation (lC5o of 0,09mM). Taken together, our findings could be helpful to further studies regarding the lectin structure and function, lectin-glycan binding characterization, as well as, the mechanisms involved in BJcuL binding to cell surface molecules, which influence cell proliferation and angiogenesi

Yuichi Endo - One of the best experts on this subject based on the ideXlab platform.

Pierre Rougé - One of the best experts on this subject based on the ideXlab platform.

  • leaves of the lamiaceae species glechoma hederacea ground ivy contain a lectin that is structurally and evolutionary related to the legume Lectins
    Plant Journal, 2003
    Co-Authors: Weifang Wang, Claire Rossi, Willy J Peumans, Paul Proost, Jianping Chen, Pierre Rougé, Els J. M. Van Damme
    Abstract:

    Summary A novel lectin has been isolated and cloned from leaves of Glechoma hederacea (ground ivy), a typical representative of the plant family Lamiaceae. Biochemical analyses indicated that the G. hederacea agglutinin (Gleheda) is a tetrameric protein consisting of four subunits pairwise linked through an interchain disulphide bridge and exhibits a preferential specificity towards N-acetylgalactosamine. Cloning of the corresponding gene and molecular modeling of the deduced sequence demonstrated that Gleheda shares high sequence similarity with the legume Lectins and exhibits the same overall fold and three-dimensional structure as the classical legume Lectins. The identification of a soluble and active legume lectin ortholog in G. hederacea not only indicates that the yet unclassified Lamiaceae Lectins belong to the same lectin family as the legume Lectins, but also sheds a new light on the specificity, physiological role and evolution of the classical legume Lectins.

  • plant Lectins a composite of several distinct families of structurally and evolutionary related proteins with diverse biological roles
    Critical Reviews in Plant Sciences, 1998
    Co-Authors: Els J. M. Van Damme, Willy J Peumans, Annick Barre, Pierre Rougé
    Abstract:

    Many plants contain carbohydrate-binding proteins that are commonly designated as Lectins, agglutinins, or hemagglutinins. Due to the obvious differences in molecular structure, biochemical properties, and carbohydrate-binding specificity, plant Lectins are usually considered a complex and heterogeneous group of proteins. Recent advances in the structural analysis of Lectins and molecular cloning of lectin genes enable subdividision of plant Lectins in a limited number of subgroups of structurally and evolutionary related proteins. Four major lectin families, namely, the legume Lectins, the chitin-binding Lectins composed of hevein domains, the type 2 ribosome-inactivating proteins, and the monocot mannose-binding Lectins comprise the majority of all currently known plant Lectins. In addition to these four large families the jacalin-related Lectins, the amaranthin family, and the Cucurbitaceae phloem Lectins are now recognized as separate subgroups. Each of the above-mentioned lectin families is discussed...

  • molecular cloning of the bark and seed Lectins from the japanese pagoda tree sophora japonica
    Plant Molecular Biology, 1997
    Co-Authors: E J M Van Damme, Pierre Rougé, Annick Barre, Willy J Peumans
    Abstract:

    cDNA clones encoding the bark and seed Lectins from Sophora japonica were isolated and their sequences analyzed. Screening of a cDNA library constructed from polyA RNA isolated from the bark resulted in the isolation of three different lectin cDNA clones. The first clone encodes the GalNAc-specific bark lectin which was originally described by Hankins et al. whereas the other clones encode the two isoforms of the mannose/glucose-specific lectin reported by Ueno et al.. Molecular cloning of the seed lectin genes revealed that Sophora seeds contain only a GalNAc-specific lectin which is highly homologous to though not identical with the GalNAc-specific lectin from the bark. All lectin polypeptides are translated from mRNAs of ca. 1.3 kb encoding a precursor carrying a signal peptide. In the case of the mannose/glucose-specific bark Lectins this precursor is post-translationally processed in two smaller peptides. Alignment of the deduced amino acid sequences of the different clones revealed striking sequence similarities between the mannose/glucose-binding and the GalNAc-specific Lectins. Furthermore, there was a high degree of sequence homology with other legume Lectins which allowed molecular modelling of the Sophora Lectins using the coordinates of the Pisum sativum, Lathyrus ochrus and Erythrina corallodendron Lectins.

Willy J Peumans - One of the best experts on this subject based on the ideXlab platform.

  • leaves of the lamiaceae species glechoma hederacea ground ivy contain a lectin that is structurally and evolutionary related to the legume Lectins
    Plant Journal, 2003
    Co-Authors: Weifang Wang, Claire Rossi, Willy J Peumans, Paul Proost, Jianping Chen, Pierre Rougé, Els J. M. Van Damme
    Abstract:

    Summary A novel lectin has been isolated and cloned from leaves of Glechoma hederacea (ground ivy), a typical representative of the plant family Lamiaceae. Biochemical analyses indicated that the G. hederacea agglutinin (Gleheda) is a tetrameric protein consisting of four subunits pairwise linked through an interchain disulphide bridge and exhibits a preferential specificity towards N-acetylgalactosamine. Cloning of the corresponding gene and molecular modeling of the deduced sequence demonstrated that Gleheda shares high sequence similarity with the legume Lectins and exhibits the same overall fold and three-dimensional structure as the classical legume Lectins. The identification of a soluble and active legume lectin ortholog in G. hederacea not only indicates that the yet unclassified Lamiaceae Lectins belong to the same lectin family as the legume Lectins, but also sheds a new light on the specificity, physiological role and evolution of the classical legume Lectins.

  • plant Lectins a composite of several distinct families of structurally and evolutionary related proteins with diverse biological roles
    Critical Reviews in Plant Sciences, 1998
    Co-Authors: Els J. M. Van Damme, Willy J Peumans, Annick Barre, Pierre Rougé
    Abstract:

    Many plants contain carbohydrate-binding proteins that are commonly designated as Lectins, agglutinins, or hemagglutinins. Due to the obvious differences in molecular structure, biochemical properties, and carbohydrate-binding specificity, plant Lectins are usually considered a complex and heterogeneous group of proteins. Recent advances in the structural analysis of Lectins and molecular cloning of lectin genes enable subdividision of plant Lectins in a limited number of subgroups of structurally and evolutionary related proteins. Four major lectin families, namely, the legume Lectins, the chitin-binding Lectins composed of hevein domains, the type 2 ribosome-inactivating proteins, and the monocot mannose-binding Lectins comprise the majority of all currently known plant Lectins. In addition to these four large families the jacalin-related Lectins, the amaranthin family, and the Cucurbitaceae phloem Lectins are now recognized as separate subgroups. Each of the above-mentioned lectin families is discussed...

  • molecular cloning of the bark and seed Lectins from the japanese pagoda tree sophora japonica
    Plant Molecular Biology, 1997
    Co-Authors: E J M Van Damme, Pierre Rougé, Annick Barre, Willy J Peumans
    Abstract:

    cDNA clones encoding the bark and seed Lectins from Sophora japonica were isolated and their sequences analyzed. Screening of a cDNA library constructed from polyA RNA isolated from the bark resulted in the isolation of three different lectin cDNA clones. The first clone encodes the GalNAc-specific bark lectin which was originally described by Hankins et al. whereas the other clones encode the two isoforms of the mannose/glucose-specific lectin reported by Ueno et al.. Molecular cloning of the seed lectin genes revealed that Sophora seeds contain only a GalNAc-specific lectin which is highly homologous to though not identical with the GalNAc-specific lectin from the bark. All lectin polypeptides are translated from mRNAs of ca. 1.3 kb encoding a precursor carrying a signal peptide. In the case of the mannose/glucose-specific bark Lectins this precursor is post-translationally processed in two smaller peptides. Alignment of the deduced amino acid sequences of the different clones revealed striking sequence similarities between the mannose/glucose-binding and the GalNAc-specific Lectins. Furthermore, there was a high degree of sequence homology with other legume Lectins which allowed molecular modelling of the Sophora Lectins using the coordinates of the Pisum sativum, Lathyrus ochrus and Erythrina corallodendron Lectins.

  • cloning and characterization of the lectin cdna clones from onion shallot and leek
    Plant Molecular Biology, 1993
    Co-Authors: E J M Van Damme, Arpad Pusztai, Irwin J. Goldstein, Koen Smeets, I Engelborghs, H Aelbers, Jan Balzarini, F Van Leuven, Willy J Peumans
    Abstract:

    Characterization of the Lectins from onion (Allium cepa), shallot (A. ascalonicum) and leek (A. porrum) has shown that these Lectins differ from previously isolated Alliaceae Lectins not only in their molecular structure but also in their ability to inhibit retrovirus infection of target cells. cDNA libraries constructed from poly(A)-rich RNA isolated from young shoots of onion, shallot and leek were screened for lectin cDNA clones using colony hybridization. Sequence analysis of the lectin cDNA clones from these three species revealed a high degree of sequence similarity both at the nucleotide and at the amino acid level. Apparently the onion, shallot and leek Lectins are translated from mRNAs of ca. 800 nucleotides. The primary translation products are preproproteins (ca. 19 kDa) which are converted into the mature lectin polypeptides (12.5-13 kDa) after post-translational modifications. Southern blot analysis of genomic DNA has shown that the Lectins are most probably encoded by a family of closely related genes which is in good agreement with the sequence heterogeneity found between different lectin cDNA clones of one species.