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Identification and Characterization of Protein Glycosylation using Specific Endo- and Exoglycosidases
Published on: December 26, 2011
Una endoglucosidasa con especificidad glicánica alternativa permite la remodelación ampliada de la glicoproteína
Jonathan J Goodfellow1, Kavitha Baruah, Keisuke Yamamoto
1Department of Chemistry, University of Oxford, Chemistry Research Laboratory, Mansfield Road, Oxford OX1 3TA, UK.
Journal of the American Chemical Society
|May 4, 2012
Resumen
Una nueva endoglucosidasa bacteriana, EndoS, ofrece una mayor selectividad para la remodelación del glicano proteico. Esta enzima procesa glicanos ligados a N de tipo complejo, abordando glicóformas de anticuerpos previamente difíciles.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Glucobiología Glucobiología.
- Enzimología Enzimología.
Sus antecedentes:
- Las endoglucosidasas de proteínas son cruciales para la modificación biocatalítica de los glicanos unidos a las proteínas.
- La selectividad limitada de las endoglucosidasas existentes dificulta la manipulación de diversos glicanos ligados a N.
- Ciertas estructuras de glicanos ligados a N siguen siendo refractarias a los métodos de procesamiento enzimático actuales.
Objetivo del estudio:
- Identificar y caracterizar nuevas endoglucosidasas con una mejor selectividad para la remodelación de proteínas glicanos.
- Evaluar la actividad complementaria de una endoglucosidasa bacteriana, EndoS, junto con enzimas existentes como EndoA y EndoH.
- Para permitir la manipulación efectiva de estructuras de glicanos ligados a N previamente intratables en proteínas, particularmente en anticuerpos.
Principales métodos:
- Aislamiento y caracterización de una endoglucosidasa bacteriana (EndoS) de Streptococcus pyogenes.
- Ensayos biocatalíticos para determinar la especificidad del sustrato de EndoS.
- Análisis comparativo de la actividad de EndoS contra endoglucosidasas conocidas (EndoA, EndoH) utilizando varios sustratos de glicanos ligados a N.
- Aplicación de EndoS para el procesamiento de glicanos ligados a N de tipo complejo, incluidos los con fucosilación del núcleo.
Principales resultados:
- EndoS exhibe una especificidad de sustrato distinta, que difiere de EndoA y EndoH.
- EndoS procesa eficazmente los glicanos ligados a N de tipo complejo, con o sin fucosilación del núcleo.
- EndoS no procesa los glicanos ligados a N de tipo oligomanosa o de tipo híbrido.
- Esta actividad selectiva amplía el conjunto de herramientas para la remodelación enzimática del glicano.
Conclusiones:
- La endoglucosidasa bacteriana EndoS proporciona una actividad enzimática complementaria para la modificación de la proteína glicano.
- EndoS permite el procesamiento de glicanos ligados a N de tipo complejo, incluidos los que se encuentran en las glicosformas de anticuerpos refractarios.
- Este descubrimiento avanza en el campo de la ingeniería de proteínas y la manipulación de glicanos.
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