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Expression and Purification of the Type I Metacaspase from a Cryptophyte Guillardia theta , GtMCA-I.
Strancar, Vida; van Midden, Katarina Petra; Klemencic, Marina; Funk, Christiane.
Afiliación
  • Strancar V; Faculty of Chemistry and Chemical Technology, Department of Chemistry and Biochemistry, University of Ljubljana, Ljubljana, Slovenia.
  • van Midden KP; Faculty of Chemistry and Chemical Technology, Department of Chemistry and Biochemistry, University of Ljubljana, Ljubljana, Slovenia.
  • Klemencic M; Faculty of Chemistry and Chemical Technology, Department of Chemistry and Biochemistry, University of Ljubljana, Ljubljana, Slovenia.
  • Funk C; Department of Chemistry, Umeå University, Umeå, Sweden.
Methods Mol Biol ; 2447: 1-11, 2022.
Article en En | MEDLINE | ID: mdl-35583768
Type I metacaspases are the most ubiquitous of the three metacaspase types and are present in representatives of prokaryotes, unicellular eukaryotes including yeasts, algae, and protozoa, as well as land plants. They are composed of two structural units: a catalytic so-called p20 domain with the His-Cys catalytic dyad and a regulatory p10 domain. Despite their structural homology to caspases, these proteases cleave their substrates after the positively charged amino acid residues at the P1 position, just like the metacaspases of type II and type III. We present a protocol for expression and purification of the only type I protease from a secondary endosymbiosis Guillardia theta , GtMCA-I by overexpression of its gene in BL21 (DE3) E. coli cells and one-day sequential purification using nickel-affinity, ion-exchange, and size-exclusion chromatography.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Caspasas / Escherichia coli Idioma: En Revista: Methods Mol Biol Asunto de la revista: BIOLOGIA MOLECULAR Año: 2022 Tipo del documento: Article País de afiliación: Eslovenia Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Caspasas / Escherichia coli Idioma: En Revista: Methods Mol Biol Asunto de la revista: BIOLOGIA MOLECULAR Año: 2022 Tipo del documento: Article País de afiliación: Eslovenia Pais de publicación: Estados Unidos