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Mémoire de Maîtrise
DOI
https://doi.org/10.11606/D.46.2019.tde-08052019-091340
Document
Auteur
Nom complet
Tamara Rezende de Azevedo
Unité de l'USP
Domain de Connaissance
Date de Soutenance
Editeur
São Paulo, 2000
Directeur
Jury
Ferreira, Clélia (Président)
Marinotti, Osvaldo
Silva, Ana Claudia Rasera da
Titre en portugais
Purificação e caracterização das β-glicosidases intestinais de Diatraea saccharalis (Lepidoptera)
Mots-clés en portugais
Bioquímica
Fisiologia animal
Insetos
Resumé en portugais
O Lepidoptera Diatraea saccharalis possui três β-glicosidases digestivas solúveis. As três enzimas foram resolvidas através de uma combinação de passos cromatográficos em colunas Econo Pac Q ( sistema de baixa pressão), Mono Q, Phenyl Superose, Superose e Mono P (FPLC). Duas delas, denominadas de E1 e E2, foram purificadas até a homogeneidade, enquanto a terceira (E3) foi semi-purificada. Dentre todas as técnicas cromatográficas testadas, colunas hidrofóbicas são as únicas capazes de resolver as três atividades de β-glicosidase. Os pesos moleculares relativos (determinados por SDS PAGE), pH ótimo e o pl foram respectivamente: E1, 58 K, 6,7, 7,5; . As três enzimas apresentam quatro subsítios de ligação de glicose e têm especificidade ampla, sendo capazes de clivar substratos sintéticos, dissacarídeos, oligossacarídeos e glicosídeos tóxicos produzidos por plantas. O principal papel fisiológico de E1 deve ser a clivagem de oligocelodextrinas derivadas da digestão de hemicelulose. E2 deve estar envolvida na hidrólise de glicolipídios, sendo a E3 a principal responsável pela digestão de dissacarídeos.
Titre en anglais
Purification and characterization of intestinal β-glycosidases of Diatraea saccharalis (Lepidoptera)
Mots-clés en anglais
Animal physiology
Biochemistry
Insects
Resumé en anglais
The Lepidoptera Diatraea saccharalis has three soluble digestive β-glycosidases. They were resolved by a combination of chromatographic steps, using Econo Pac Q (low-pressure system), Mono Q , Phenyl Superose, Superose 12 and Mono P (FPLC system) columns. Two enzymes called E1 and E2 were purified to homogeneity. The third (E3) was semi-purified. Hydrophobic columns are the only ones able to resolve the three β-glycosidase activities. The relative molecular weights (SDS PAGE}, pH optimum and pi values were, respectively: E1 , 58 K, 6.7, 7.5; E2, 61 K, 6.3, 7.4; E3, 61 K, 7.2, 7.4. The enzymes have four sub-sites for glucon binding in their active sites. They have broad substrate specificity to hydrolyze synthetic substrates, disaccharides, oligosaccharides and plant toxic glucosides. E1 may have the physiological role of hydrolyzing oligosaccharides derived from hemicellulose digestion. E2 may be responsible for glycolipid digestion and E3 seems to be the main enzyme that digests disaccharides.
 
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Date de Publication
2019-05-08
 
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