Expression and characterization of a recombinant endoglucanase from western corn rootworm, in Pichia pastoris.
J Insect Sci
; 14: 242, 2014.
Article
em En
| MEDLINE
| ID: mdl-25434035
The endoglucanase cDNA, Dvv-ENGase I, from western corn rootworm, Diabrotica virgifera virgifera LeConte was expressed using the GS115 methylotrophic strain of Pichia pastoris. The Dvv-ENGase I gene was cloned into the integrative plasmid pPICZαA under the control of AOX1, which is a methanol-inducible promoter. Positive clones were selected for their ability to produce the recombinant endoglucanase upon continuous methanol induction. The secreted recombinant insect endoglucanase Dvv-ENGase I has an apparent molecular mass of 29 kDa. The recombinant endo-1,4-ß-glucanase (ENGase) was able to digest the substrates: hydroxyethyl cellulose (HEC), carboxymethyl cellulose (CMC), and Whatman No. 1 filter paper. A higher accumulation of reducing sugar was evident when the P. pastoris expression medium contained HEC (1%) instead of CMC (1%). An enzymatic activity band was detected after performing electrophoretic separation under nondenaturing conditions. The biological activity of the recombinant Dvv-ENGase I was influenced by the presence of protease inhibitors in the culture medium.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Pichia
/
Besouros
/
Celulase
/
Proteínas de Insetos
Limite:
Animals
Idioma:
En
Revista:
J Insect Sci
Assunto da revista:
BIOLOGIA
Ano de publicação:
2014
Tipo de documento:
Article
País de afiliação:
Colômbia
País de publicação:
Estados Unidos