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Explorando la catálisis de la nucleósida hidrolasa en silico: estudio de dinámica molecular del sustrato unido a la
Devleena Mazumder1, Thomas C Bruice
1Department of Chemistry and Biochemistry, University of California, Santa Barbara, Santa Barbara, California 93106, USA.
Journal of the American Chemical Society
|December 6, 2002
Resumen
Las simulaciones de dinámica molecular revelan que Tyr229, no His241, actúa como la fuente de protones para la inosina-uridina nucleósido hidrolasa. La mutación de His241Ala interrumpe esta red clave de enlaces de hidrógeno, reduciendo la actividad de la enzima.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Enzimología Enzimología.
- Biología computacional Biología computacional.
Sus antecedentes:
- La enzima hidrolasa nucleosídica inosina-uridina juega un papel crucial en el metabolismo de los nucleósidos.
- Comprender su mecanismo de acción es vital para el desarrollo de fármacos y la investigación bioquímica.
- Estudios anteriores sugirieron His241 como un residuo catalítico clave.
Objetivo del estudio:
- Para aclarar el mecanismo detallado de la inosina-uridina nucleósido hidrolasa.
- Para investigar el papel específico de His241 en la actividad catalítica de la enzima.
- Para explorar la función de Tyr229 y la red de enlace de hidrógeno del sitio activo.
Principales métodos:
- Se emplearon simulaciones de dinámica molecular a largo plazo (DM).
- Las simulaciones se llevaron a cabo utilizando el agua TIP3P y las condiciones de límite estocástico.
- Se modelaron cinco complejos distintos de enzima-sustrato y de estado de transición de la enzima, incluido un mutante His241Ala.
Principales resultados:
- Las simulaciones de dinámica molecular indican que Tyr229, no His241, actúa como la fuente de protones de la base nuclear saliente.
- La mutación His241Ala reduce significativamente la actividad enzimática al interrumpir la red de enlace de hidrógeno del sitio activo que involucra a Tyr229.
- La protonación de His241 no alteró las estructuras complejas de la enzima, un hallazgo inesperado.
- La enzima exhibe una dependencia del pH en forma de campana, relacionada con la disociación Asp10/Asp15 y los estados de protonación Tyr229.
- La estabilización del intermediario de iones de ribooxocarbenio involucra Asn168 y Asp14, con Asp14 enlazado con hidrógeno a la ribosa 2'-OH durante el 96% del tiempo de simulación.
- Un mecanismo de lanzadera de protones que involucra agua y residuos de aspartato (Asp10, Asp15) puede ayudar al ataque nucleofílico por agua.
Conclusiones:
- Tyr229 es el donante primario de protones en el mecanismo catalítico de la inosina-uridina nucleósido hidrolasa.
- La integridad de la red de enlaces de hidrógeno, particularmente la que involucra a Tyr229, es esencial para la función de la enzima.
- Investigaciones adicionales sobre el movimiento anticorrelacionado entre Tyr229 y Asn168 pueden revelar ideas sobre las conformaciones de ataque cercano.
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