Show simple item record

dc.date.accessioned 2018-05-10T15:59:56Z
dc.date.available 2018-05-10T15:59:56Z
dc.identifier.uri http://digital.cic.gba.gob.ar/handle/11746/7254
dc.title High-level expression of Falcipain-2 in Escherichia coli by codon optimization and auto-induction en
dc.type Artículo es
dcterms.abstract Falcipain-2, the major cysteine hemoglobinase from the human malaria parasite Plasmodium falciparum, is critical for parasite development and is considered a promising chemotherapeutic target. In order to facilitate the high-throughput screening of Falcipain-2 inhibitors from natural sources, we developed an economic and highly-productive overexpression system in Escherichia coli using a codon-optimized proFalcipain-2 construct. Very high expression levels (35–55% of total host proteins) were observed when proFalcaipain-2 expression was induced with 1 mM isopropyl-1-thio-b-D-galactopyranoside (IPTG) in several E. coli strains, with the highest level observed for BL21(DE3). A lower expression ( 40% of total host proteins) was observed when BL21(DE3) was grown in ZYM-5052 auto-induction medium, containing 0.2% lactose as inducer. However, the culture grew to notably higher cellular density, increasing 1.5 times the overall yield of the system when compared with conventional IPTG-induction. Although several conditions were modified to achieve the expression of soluble and active Falcipain-2, the enzyme was mainly obtained in the form of insoluble aggregates. After purification and refolding, 50 mg of active enzyme were obtained per liter of culture at low cost using a regular incubator shaker, and recombinant Falcipain-2 exhibited structural and functional characteristics very similar to the natural counterpart. Due to its versatility and simplicity, this strategy can be straightforwardly adapted to other proteins from Plasmodium species or any other organism with an AT-rich genome. en
dcterms.extent p. 59–69 es
dcterms.issued 2012-05
dcterms.language Inglés es
dcterms.license Attribution-NonCommercial-ShareAlike 4.0 International (BY-NC-SA 4.0) es
dcterms.publisher Elsevier es
dcterms.subject Falcipain-2 en
dcterms.subject Plasmodium falciparum es
dcterms.subject Synthetic gene en
dcterms.subject Codon optimization en
dcterms.subject Auto-induction en
cic.version info:eu-repo/semantics/publishedVersion es
dcterms.creator.author Salas Sarduy, Emir es
dcterms.creator.author Cabrera Muñoz, Aymara es
dcterms.creator.author Trejo, Sebastián Alejandro es
dcterms.creator.author Chavéz Planes, María de los A. es
cic.lugarDesarrollo Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús es
dcterms.subject.materia Bioquímica y Biología Molecular es
dcterms.identifier.url Recurso online es
dcterms.identifier.other doi:10.1016/j.pep.2012.03.008 es
dcterms.isPartOf.issue vol. 83, no. 1 es
dcterms.isPartOf.series Protein Expression and Purification es
cic.isPeerReviewed true es
cic.isFulltext true es


Files

  • Icon

    Documento completo 

    PDF file (1.697Mb)

  • This item appears in the following Collection(s)

    Show simple item record