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Characterization of soluble CD39 (SolCD39/NTPDase1) from PiggyBac nonviral system as a tool to control the nucleotides level.
Beckenkamp, Liziane Raquel; Iser, Isabele Cristiana; Onzi, Giovana Ravizzoni; Fontoura, Dieine Maira Soares da; Bertoni, Ana Paula Santin; Sévigny, Jean; Lenz, Guido; Wink, Márcia Rosângela.
Afiliación
  • Beckenkamp LR; Laboratory of Cell Biology, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, Brazil.
  • Iser IC; Laboratory of Cell Biology, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, Brazil.
  • Onzi GR; Department of Biophysics and Center of Biotechnology, Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, RS, Brazil.
  • Fontoura DMSD; Laboratory of Cell Biology, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, Brazil.
  • Bertoni APS; Laboratory of Cell Biology, Universidade Federal de Ciências da Saúde de Porto Alegre, Porto Alegre, Brazil.
  • Sévigny J; Département de Microbiologie-Infectiologie et d'Immunologie, Faculté de Médecine, Université Laval, Québec City, QC G1V 0A6, Canada.
  • Lenz G; Centre de recherche du CHU de Québec-Université Laval, Québec City, QC G1V 4G2, Canada.
  • Wink MR; Department of Biophysics and Center of Biotechnology, Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, RS, Brazil.
Biochem J ; 476(11): 1637-1651, 2019 06 11.
Article en En | MEDLINE | ID: mdl-31085558
Extracellular ATP (eATP) and its metabolites have emerged as key modulators of different diseases and comprise a complex pathway called purinergic signaling. An increased number of tools have been developed to study the role of nucleotides and nucleosides in cell proliferation and migration, influence on the immune system and tumor progression. These tools include receptor agonists/antagonists, engineered ectonucleotidases, interference RNAs and ectonucleotidase inhibitors that allow the control and quantification of nucleotide levels. NTPDase1 (also called apyrase, ecto-ATPase and CD39) is one of the main enzymes responsible for the hydrolysis of eATP, and purified enzymes, such as apyrase purified from potato, or engineered as soluble CD39 (SolCD39), have been widely used in in vitro and in vivo experiments. However, the commercial apyrase had its effects recently questioned and SolCD39 exhibits limitations, such as short half-life and need of high doses to reach the expected enzymatic activity. Therefore, this study investigated a non-viral method to improve the overexpression of SolCD39 and evaluated its impact on other enzymes of the purinergic system. Our data demonstrated that PiggyBac transposon system proved to be a fast and efficient method to generate cells stably expressing SolCD39, producing high amounts of the enzyme from a limited number of cells and with high hydrolytic activity. In addition, the soluble form of NTPDase1/CD39 did not alter the expression or catalytic activity of other enzymes from the purinergic system. Altogether, these findings set the groundwork for prospective studies on the function and therapeutic role of eATP and its metabolites in physiological and pathological conditions.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Apirasa / Antígenos CD Tipo de estudio: Observational_studies Límite: Animals Idioma: En Revista: Biochem J Año: 2019 Tipo del documento: Article País de afiliación: Brasil Pais de publicación: Reino Unido
Texto completo
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Imprimir
XML
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Apirasa / Antígenos CD Tipo de estudio: Observational_studies Límite: Animals Idioma: En Revista: Biochem J Año: 2019 Tipo del documento: Article País de afiliación: Brasil Pais de publicación: Reino Unido