Navegando por Palavras-chave "protein structure"

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    Biochemical and biophysical properties of a highly active recombinant arginase from Leishmania (Leishmania) amazonensis and subcellular localization of native enzyme
    (Elsevier B.V., 2008-06-01) Silva, Edson Roberto da; Laranjeira da Silva, Maria Fernanda; Fischer, Hannes; Mortara, Renato A. [UNIFESP]; Mayer, Mario Gustavo; Framesqui, Karine; Silber, Ariel Mariano; Floeter-Winter, Lucile Maria; Universidade de São Paulo (USP); Univ Luterana Brasil; Universidade Federal de São Paulo (UNIFESP); Inst Butantan Serv Genet
    Arginase (L-arginine amidinohydrolase, E.C. 3.5.3.1) is a metalloenzyme that catalyses the hydrolysis of L-arginine to L-ornithine and urea. in Leishmania spp., the biological role of the enzyme may be involved in modulating NO production upon macrophage infection. Previously, we cloned and characterized the arginase gene from Leishmania (Leishmania) amazonensis. in the present work, we successfully expressed the recombinant enzyme in E. coli and performed biochemical and biophysical characterization of both the native and recombinant enzymes. We obtained K-M and V-max. values of 23.9(+/- 0.96) mM and 192.3 mu mol/min mg protein (+/- 14.3), respectively, for the native enzyme. for the recombinant counterpart, K-M was 21.5(+/- 0.90) mM and V-max was 144.9(+/- 8.9) mu mol/min mg. Antibody against the recombinant protein confirmed a glycosomal cellular localization of the enzyme in promastigotes. Data from light scattering and small angle X-ray scattering showed that a trimeric state is the active form of the protein. We determined empirically that a manganese wash at room temperature is the best condition to purify active enzyme. the interaction of the recombinant protein with the immobilized nickel also allowed us to confirm the structural disposition of histidine at positions 3 and 324. the determined structural parameters provide substantial data to facilitate the search for selective inhibitors of parasitic sources of arginase, which could subsequently point to a candidate for leishmaniasis therapy. (c) 2008 Elsevier B.V. All rights reserved.
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    The structural molecular biology network of the State of São Paulo, Brazil
    (Academia Brasileira de Ciências, 2006-06-01) Barbosa, João A.r.g.; Netto, Luis E.s.; Farah, Chuck S.; Schenkman, Sergio [UNIFESP]; Meneghini, Rogério; Centro de Biologia Molecular Estrutural Laboratório Nacional de Luz Síncrotron; Universidade de São Paulo (USP); Universidade Federal de São Paulo (UNIFESP); Centro Latino-Americano e do Caribe de Informação em Ciências da Saúde BIREME
    This article describes the achievements of the Structural Molecular Biology Network (SMolBNet), a collaborative program of structural molecular biology, centered in the State of São Paulo, Brazil, and supported by São Paulo State Funding Agency (FAPESP). It gathers twenty scientific groups and is coordinated by the scientific staff of the Center of Structural Molecular Biology, at the National Laboratory of Synchrotron Light (LNLS), in Campinas. The SMolBNet program has been aimed at 1) solving the structure of proteins of interest related to the research projects of the groups. In some cases, the choice has been to select proteins of unknown function or of possible novel structure obtained from the sequenced genomes of the FAPESP genomic program; 2) providing the groups with training in all the steps of the protein structure determination: gene cloning, protein expression, protein purification, protein crystallization and structure determination. Having begun in 2001, the program has been successful in both aims. Here, four groups reveal their participation in the program and describe the structural aspects of the proteins they have selected to study.