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Catálisis ácido-base general mediada por nucleobases en la ribozima de la horquilla del cabello
Stephanie Kath-Schorr1, Timothy J Wilson, Nan-Sheng Li
1Cancer Research UK, Nucleic Acid Structure Research Group, MSI/WTB Complex, The University of Dundee, Dow Street, Dundee DD1 5EH, UK.
Journal of the American Chemical Society
|September 11, 2012
Resumen
La horquilla es la ribozima ribozima.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Biología Molecular Biología Molecular
- La catálisis del ARN.
Sus antecedentes:
- El mecanismo catalítico de la división/ligación del ARN mediada por la ribozima de la horquilla no se entiende completamente.
- La evidencia experimental sugiere roles para la adenina (A38) y la guanina (G8) en la catálisis.
- Las hipótesis anteriores proponían la catálisis ácido-base general por estos residuos.
Objetivo del estudio:
- Para dilucidar el mecanismo catalítico de la ribozima de la horquilla.
- Para investigar las funciones de la adenina (A38) y la guanina (G8) en la escisión del enlace fosfodiéster de ARN.
- Para determinar si A38 y G8 funcionan en la catálisis ácido-base general.
Principales métodos:
- Mutagénesis dirigida al sitio de los residuos de ribozima A38 y G8 de la horquilla del cabello.
- Síntesis de sustratos de ARN con un grupo de salida de 5'-pro-tiosulfato (5'-PS).
- Análisis cinético de las tasas de escisión en un rango de valores de pH.
Principales resultados:
- La sustitución de A38 por purina (A38P) redujo la tasa de escisión, que fue revertida por la modificación 5'-PS.
- La velocidad de escisión del sustrato 5'-PS por la ribozima A38P mostró una dependencia del pH consistente con un catalizador ácido general.
- La sustitución de G8 por diaminopurina dio lugar a tasas de escisión dependientes del pH indicativas de un catalizador de base general.
Conclusiones:
- Los datos experimentales apoyan fuertemente un mecanismo general de catálisis ácido-base para la ribozima de horquilla.
- La adenina (A38) probablemente actúa como un catalizador ácido general.
- La guanina (G8) probablemente actúa como un catalizador de base general en el mecanismo de la ribozima de la horquilla.
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