The Experts below are selected from a list of 2313 Experts worldwide ranked by ideXlab platform

Marc Fransen - One of the best experts on this subject based on the ideXlab platform.

  • 2,4-dienoyl-CoA reductase
    2013
    Co-Authors: Marc Fransen, Paul P Van Veldhoven, Suresh Subramani
    Abstract:

    To elucidate unknown mammalian peroxisomal enzymes and functions, we subjected M13 phage expressing fusions between the gene encoding protein VI and a rat liver cDNA library to an immunoaffinity selection process in �itro (Biopanning) with the use of antibodies raised against peroxisomal subfractions. In an initial series of Biopanning experiments, four different cDNA clones were obtained. These cDNA species encoded two previously identified peroxisomal enzymes, catalase and urate oxidase, and two novel proteins that contained a C-terminal peroxisomal targeting signal (PTS1). A primary structure analysis of these novel proteins revealed that one, ending in the tripeptide AKL, is homologous to the yeast peroxisomal 2,4-dienoyl-CoA reductase (EC 1.3.1.34; DCR), an enzyme required for the degradation of unsaturated fatty acids, and that the other, ending in the tripeptide SRL, is a putative member of the shortchain dehydrogenase�reductase (SDR) family, with three isoforms. Green fluorescent protein (GFP) fusions encoding GFP– DCR-AKL, GFP–DCR, GFP–SDR-SRL and GFP–SDR were expressed in mammalian cells. The analysis of the subcellular location of the recombinant fusion proteins confirmed the peroxisomal localization of GFP–DCR-AKL and GFP–SDR-SRL, as well as the functionality of the PTS1. That the AKL protein is indeed an NADPH-dependent DCR was demonstrated by showing DCR activity of the bacterially expressed protein. These results demonstrate at the molecular level that mammalian peroxisomes do indeed contain a DCR. In addition, the results presented here indicate that the protein VI display system is suitable for the isolation of rare cDNA clones from cDNA libraries and that this technology facilitates the identification of novel peroxisomal proteins. Key words: fatty acid oxidation, M13 phage, peroxisomal metabolism

  • identification of peroxisomal proteins by using m13 phage protein vi phage display molecular evidence that mammalian peroxisomes contain a 2 4 dienoyl coa reductase
    Biochemical Journal, 1999
    Co-Authors: Marc Fransen, Paul P Van Veldhoven, Suresh Subramani
    Abstract:

    To elucidate unknown mammalian peroxisomal enzymes and functions, we subjected M13 phage expressing fusions between the gene encoding protein VI and a rat liver cDNA library to an immunoaffinity selection process in vitro (Biopanning) with the use of antibodies raised against peroxisomal subfractions. In an initial series of Biopanning experiments, four different cDNA clones were obtained. These cDNA species encoded two previously identified peroxisomal enzymes, catalase and urate oxidase, and two novel proteins that contained a C-terminal peroxisomal targeting signal (PTS1). A primary structure analysis of these novel proteins revealed that one, ending in the tripeptide AKL, is homologous to the yeast peroxisomal 2,4-dienoyl-CoA reductase (EC 1.3.1.34; DCR), an enzyme required for the degradation of unsaturated fatty acids, and that the other, ending in the tripeptide SRL, is a putative member of the short-chain dehydrogenase/reductase (SDR) family, with three isoforms. Green fluorescent protein (GFP) fusions encoding GFP-DCR-AKL, GFP-DCR, GFP-SDR-SRL and GFP-SDR were expressed in mammalian cells. The analysis of the subcellular location of the recombinant fusion proteins confirmed the peroxisomal localization of GFP-DCR-AKL and GFP-SDR-SRL, as well as the functionality of the PTS1. That the AKL protein is indeed an NADPH-dependent DCR was demonstrated by showing DCR activity of the bacterially expressed protein. These results demonstrate at the molecular level that mammalian peroxisomes do indeed contain a DCR. In addition, the results presented here indicate that the protein VI display system is suitable for the isolation of rare cDNA clones from cDNA libraries and that this technology facilitates the identification of novel peroxisomal proteins.

Leonard W Seymour - One of the best experts on this subject based on the ideXlab platform.

  • bacteriophage Biopanning in human tumour biopsies to identify cancer specific targeting ligands
    Journal of Drug Targeting, 2007
    Co-Authors: Fukuto Maruta, D J Kerr, Leonard W Seymour, Noriyuki Akita, Jun Nakayama, Shinichi Miyagawa, Tariq Ismail, D C Rowlands, Kerry D Fisher, Alan L Parker
    Abstract:

    Intravenous targeting of anticancer agents should improve both efficacy and therapeutic index. However, rational design of targeting constructs requires detailed definition of receptor targets and must take account of polarised tissue architecture that may restrict access to chosen receptors from the bloodstream. Bacteriophage Biopanning provides a solution to this problem, identifying targeting sequences by functional selection rather than design, although reiterative panning in polarized human tumours has not previously been attempted. Here, we report an ex vivo, intra-arterial method for Biopanning in freshly-resected human tumours, enabling reiterative selection of oligopeptide sequences capable of intravascular targeting to human colorectal tumours. Significant consensus was observed after two rounds of panning in tumours from different patients, and lead sequences demonstrated tumour targeting in samples from unrelated patients. This novel approach may be applicable to a wide range of settings, thus enabling iteration of consensus targeting sequences for tumour imaging and selective delivery of anticancer agents.

  • use of a phage display library to identify oligopeptides binding to the lumenal surface of polarized endothelium by ex vivo perfusion of human umbilical veins
    Journal of Drug Targeting, 2003
    Co-Authors: Fukuto Maruta, Alan L Parker, Kerry Fisher, D J Kerr, Paul G Murray, Leonard W Seymour
    Abstract:

    Human endothelial-specific targeting peptides were identified by Biopanning within freshly-obtained human umbilical cords. Umbilical veins were cleaned in situ and M13 phage display libraries were passed through the cords. Tightly bound phage were recovered following isolation of endothelial cells by collagenase digestion and homogenisation, allowing production of enriched phage libraries for subsequent rounds of panning. After five rounds of Biopanning, five promising sequences were selected and the binding of the corresponding phage clones was compared in perfused umbilical veins. Each of these peptides showed substantial binding, although the clone encoding the heptapeptide KPSGLTY showed the greatest, some 89-times greater than insertless phage. Binding of this phage clone was examined to cells in vitro, where it demonstrated at least five-times greater binding to isolated human umbilical vein endothelial cells than to 911, SKOV3, B16F10 and Cos7 cells. These initial peptides may prove useful targeting agents for endothelial-selective delivery, and this powerful approach should be readily applicable to Biopanning in a broad range of human vessels ex vivo.

  • use of a phage display library to identify oligopeptides binding to the lumenal surface of polarized endothelium by ex vivo perfusion of human umbilical veins
    Journal of Drug Targeting, 2003
    Co-Authors: Fukuto Maruta, Alan L Parker, Kerry Fisher, D J Kerr, Paul G Murray, Leonard W Seymour
    Abstract:

    Human endothelial-specific targeting peptides were identified by Biopanning within freshly-obtained human umbilical cords. Umbilical veins were cleaned in situ and M13 phage display libraries were passed through the cords. Tightly bound phage were recovered following isolation of endothelial cells by collagenase digestion and homogenisation, allowing production of enriched phage libraries for subsequent rounds of panning. After five rounds of Biopanning, five promising sequences were selected and the binding of the corresponding phage clones was compared in perfused umbilical veins. Each of these peptides showed substantial binding, although the clone encoding the heptapeptide KPSGLTY showed the greatest, some 89-times greater than insertless phage. Binding of this phage clone was examined to cells in vitro, where it demonstrated at least five-times greater binding to isolated human umbilical vein endothelial cells than to 911, SKOV3, B16F10 and Cos7 cells. These initial peptides may prove useful targeting agents for endothelial-selective delivery, and this powerful approach should be readily applicable to Biopanning in a broad range of human vessels ex vivo.

Suresh Subramani - One of the best experts on this subject based on the ideXlab platform.

  • 2,4-dienoyl-CoA reductase
    2013
    Co-Authors: Marc Fransen, Paul P Van Veldhoven, Suresh Subramani
    Abstract:

    To elucidate unknown mammalian peroxisomal enzymes and functions, we subjected M13 phage expressing fusions between the gene encoding protein VI and a rat liver cDNA library to an immunoaffinity selection process in �itro (Biopanning) with the use of antibodies raised against peroxisomal subfractions. In an initial series of Biopanning experiments, four different cDNA clones were obtained. These cDNA species encoded two previously identified peroxisomal enzymes, catalase and urate oxidase, and two novel proteins that contained a C-terminal peroxisomal targeting signal (PTS1). A primary structure analysis of these novel proteins revealed that one, ending in the tripeptide AKL, is homologous to the yeast peroxisomal 2,4-dienoyl-CoA reductase (EC 1.3.1.34; DCR), an enzyme required for the degradation of unsaturated fatty acids, and that the other, ending in the tripeptide SRL, is a putative member of the shortchain dehydrogenase�reductase (SDR) family, with three isoforms. Green fluorescent protein (GFP) fusions encoding GFP– DCR-AKL, GFP–DCR, GFP–SDR-SRL and GFP–SDR were expressed in mammalian cells. The analysis of the subcellular location of the recombinant fusion proteins confirmed the peroxisomal localization of GFP–DCR-AKL and GFP–SDR-SRL, as well as the functionality of the PTS1. That the AKL protein is indeed an NADPH-dependent DCR was demonstrated by showing DCR activity of the bacterially expressed protein. These results demonstrate at the molecular level that mammalian peroxisomes do indeed contain a DCR. In addition, the results presented here indicate that the protein VI display system is suitable for the isolation of rare cDNA clones from cDNA libraries and that this technology facilitates the identification of novel peroxisomal proteins. Key words: fatty acid oxidation, M13 phage, peroxisomal metabolism

  • identification of peroxisomal proteins by using m13 phage protein vi phage display molecular evidence that mammalian peroxisomes contain a 2 4 dienoyl coa reductase
    Biochemical Journal, 1999
    Co-Authors: Marc Fransen, Paul P Van Veldhoven, Suresh Subramani
    Abstract:

    To elucidate unknown mammalian peroxisomal enzymes and functions, we subjected M13 phage expressing fusions between the gene encoding protein VI and a rat liver cDNA library to an immunoaffinity selection process in vitro (Biopanning) with the use of antibodies raised against peroxisomal subfractions. In an initial series of Biopanning experiments, four different cDNA clones were obtained. These cDNA species encoded two previously identified peroxisomal enzymes, catalase and urate oxidase, and two novel proteins that contained a C-terminal peroxisomal targeting signal (PTS1). A primary structure analysis of these novel proteins revealed that one, ending in the tripeptide AKL, is homologous to the yeast peroxisomal 2,4-dienoyl-CoA reductase (EC 1.3.1.34; DCR), an enzyme required for the degradation of unsaturated fatty acids, and that the other, ending in the tripeptide SRL, is a putative member of the short-chain dehydrogenase/reductase (SDR) family, with three isoforms. Green fluorescent protein (GFP) fusions encoding GFP-DCR-AKL, GFP-DCR, GFP-SDR-SRL and GFP-SDR were expressed in mammalian cells. The analysis of the subcellular location of the recombinant fusion proteins confirmed the peroxisomal localization of GFP-DCR-AKL and GFP-SDR-SRL, as well as the functionality of the PTS1. That the AKL protein is indeed an NADPH-dependent DCR was demonstrated by showing DCR activity of the bacterially expressed protein. These results demonstrate at the molecular level that mammalian peroxisomes do indeed contain a DCR. In addition, the results presented here indicate that the protein VI display system is suitable for the isolation of rare cDNA clones from cDNA libraries and that this technology facilitates the identification of novel peroxisomal proteins.

Fukuto Maruta - One of the best experts on this subject based on the ideXlab platform.

  • bacteriophage Biopanning in human tumour biopsies to identify cancer specific targeting ligands
    Journal of Drug Targeting, 2007
    Co-Authors: Fukuto Maruta, D J Kerr, Leonard W Seymour, Noriyuki Akita, Jun Nakayama, Shinichi Miyagawa, Tariq Ismail, D C Rowlands, Kerry D Fisher, Alan L Parker
    Abstract:

    Intravenous targeting of anticancer agents should improve both efficacy and therapeutic index. However, rational design of targeting constructs requires detailed definition of receptor targets and must take account of polarised tissue architecture that may restrict access to chosen receptors from the bloodstream. Bacteriophage Biopanning provides a solution to this problem, identifying targeting sequences by functional selection rather than design, although reiterative panning in polarized human tumours has not previously been attempted. Here, we report an ex vivo, intra-arterial method for Biopanning in freshly-resected human tumours, enabling reiterative selection of oligopeptide sequences capable of intravascular targeting to human colorectal tumours. Significant consensus was observed after two rounds of panning in tumours from different patients, and lead sequences demonstrated tumour targeting in samples from unrelated patients. This novel approach may be applicable to a wide range of settings, thus enabling iteration of consensus targeting sequences for tumour imaging and selective delivery of anticancer agents.

  • use of a phage display library to identify oligopeptides binding to the lumenal surface of polarized endothelium by ex vivo perfusion of human umbilical veins
    Journal of Drug Targeting, 2003
    Co-Authors: Fukuto Maruta, Alan L Parker, Kerry Fisher, D J Kerr, Paul G Murray, Leonard W Seymour
    Abstract:

    Human endothelial-specific targeting peptides were identified by Biopanning within freshly-obtained human umbilical cords. Umbilical veins were cleaned in situ and M13 phage display libraries were passed through the cords. Tightly bound phage were recovered following isolation of endothelial cells by collagenase digestion and homogenisation, allowing production of enriched phage libraries for subsequent rounds of panning. After five rounds of Biopanning, five promising sequences were selected and the binding of the corresponding phage clones was compared in perfused umbilical veins. Each of these peptides showed substantial binding, although the clone encoding the heptapeptide KPSGLTY showed the greatest, some 89-times greater than insertless phage. Binding of this phage clone was examined to cells in vitro, where it demonstrated at least five-times greater binding to isolated human umbilical vein endothelial cells than to 911, SKOV3, B16F10 and Cos7 cells. These initial peptides may prove useful targeting agents for endothelial-selective delivery, and this powerful approach should be readily applicable to Biopanning in a broad range of human vessels ex vivo.

  • use of a phage display library to identify oligopeptides binding to the lumenal surface of polarized endothelium by ex vivo perfusion of human umbilical veins
    Journal of Drug Targeting, 2003
    Co-Authors: Fukuto Maruta, Alan L Parker, Kerry Fisher, D J Kerr, Paul G Murray, Leonard W Seymour
    Abstract:

    Human endothelial-specific targeting peptides were identified by Biopanning within freshly-obtained human umbilical cords. Umbilical veins were cleaned in situ and M13 phage display libraries were passed through the cords. Tightly bound phage were recovered following isolation of endothelial cells by collagenase digestion and homogenisation, allowing production of enriched phage libraries for subsequent rounds of panning. After five rounds of Biopanning, five promising sequences were selected and the binding of the corresponding phage clones was compared in perfused umbilical veins. Each of these peptides showed substantial binding, although the clone encoding the heptapeptide KPSGLTY showed the greatest, some 89-times greater than insertless phage. Binding of this phage clone was examined to cells in vitro, where it demonstrated at least five-times greater binding to isolated human umbilical vein endothelial cells than to 911, SKOV3, B16F10 and Cos7 cells. These initial peptides may prove useful targeting agents for endothelial-selective delivery, and this powerful approach should be readily applicable to Biopanning in a broad range of human vessels ex vivo.

Moon Young Yang - One of the best experts on this subject based on the ideXlab platform.

  • gold binding peptide identified from microfluidic Biopanning an experimental and molecular dynamics study
    Langmuir, 2019
    Co-Authors: Dong Jae Lee, Hyun Su Park, Kunmo Koo, Jeong Yong Lee, Yoon Sung Nam, Wonhee Lee, Moon Young Yang
    Abstract:

    Biopanning refers to the processes of screening peptides with a high affinity to a target material. Microfluidic Biopanning has advantages compared to conventional Biopanning which requires large amounts of the target material and involves inefficient multiple pipetting steps to remove nonspecific or low-affinity peptides. Here, we fabricate a microfluidic Biopanning system to identify a new gold-binding peptide (GBP). A polydimethylsiloxane microfluidic device is fabricated and bonded to a glass slide with a gold pattern that is deposited by electron-beam evaporation. The microfluidic Biopanning system can provide high adjustability in the washing step during the Biopanning process because the liquid flow rate and the resulting shear stress can be precisely controlled. The surface plasmon resonance analysis shows that the binding affinity of the identified GBP is comparable to previously reported GBPs. Moreover, molecular dynamics simulations are performed to understand its binding affinity against the g...

  • Gold Binding Peptide Identified from Microfluidic Biopanning: An Experimental and Molecular Dynamics Study
    AMER CHEMICAL SOC, 2019
    Co-Authors: Dong Jae Lee, Hyun Su Park, Kunmo Koo, Jeong Yong Lee, Yoon Sung Nam, Wonhee Lee, Moon Young Yang
    Abstract:

    Biopanning refers to the processes of screening peptides with a high affinity to a target material. Microfluidic Biopanning has advantages compared to conventional Biopanning which requires large amounts of the target material and involves inefficient multiple pipetting steps to remove nonspecific or low-affinity peptides. Here, we fabricate a microfluidic Biopanning system to identify a new gold-binding peptide (GBP). A polydimethylsiloxane microfluidic device is fabricated and bonded to a glass slide with a gold pattern that is deposited by electron-beam evaporation. The microfluidic Biopanning system can provide high adjustability in the washing step during the Biopanning process because the liquid flow rate and the resulting shear stress can be precisely controlled. The surface plasmon resonance analysis shows that the binding affinity of the identified GBP is comparable to previously reported GBPs. Moreover, molecular dynamics simulations are performed to understand its binding affinity against the gold surface in detail. Theoretical calculations suggest that the association and dissociation rates of the GBPs depend on their sequence-dependent conformations and interactions with the gold surface. These findings provide insight into designing efficient Biopanning tools and peptides with a high affinity for various target materials. © Copyright © 2018 American Chemical Societ

  • Gold Binding Peptide Identified from Microfluidic Biopanning: An Experimental and Molecular Dynamics Study
    2018
    Co-Authors: Dong Jae Lee, Hyun Su Park, Kunmo Koo, Jeong Yong Lee, Yoon Sung Nam, Wonhee Lee, Moon Young Yang
    Abstract:

    Biopanning refers to the processes of screening peptides with a high affinity to a target material. Microfluidic Biopanning has advantages compared to conventional Biopanning which requires large amounts of the target material and involves inefficient multiple pipetting steps to remove nonspecific or low-affinity peptides. Here, we fabricate a microfluidic Biopanning system to identify a new gold-binding peptide (GBP). A polydimethylsiloxane microfluidic device is fabricated and bonded to a glass slide with a gold pattern that is deposited by electron-beam evaporation. The microfluidic Biopanning system can provide high adjustability in the washing step during the Biopanning process because the liquid flow rate and the resulting shear stress can be precisely controlled. The surface plasmon resonance analysis shows that the binding affinity of the identified GBP is comparable to previously reported GBPs. Moreover, molecular dynamics simulations are performed to understand its binding affinity against the gold surface in detail. Theoretical calculations suggest that the association and dissociation rates of the GBPs depend on their sequence-dependent conformations and interactions with the gold surface. These findings provide insight into designing efficient Biopanning tools and peptides with a high affinity for various target materials