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La unión al sustrato regula la señalización redox en la primasa del ADN humano
Elizabeth O'Brien1, Marilyn E Holt2, Lauren E Salay2
1Division of Chemistry and Chemical Engineering , California Institute of Technology , Pasadena , California 91125 , United States.
Journal of the American Chemical Society
|November 16, 2018
Resumen
La ADN primasa utiliza un grupo [4Fe4S] para la señalización durante la replicación del ADN. Su potencial redox cambia al rango fisiológico al unirse al ADN y a las NTP, aumentando la actividad enzimática.
Área de la Ciencia:
- La bioquímica
- Biología molecular
- Enzimología
Sus antecedentes:
- La replicación del ADN requiere de la ADN primasa para sintetizar los primers de ARN.
- La primasa es un heterodímero (p48/p58) con dominios funcionales distintos.
- El grupo [4Fe4S] en p58 es crucial para la actividad de la enzima.
Objetivo del estudio:
- Para investigar las propiedades de conmutación redox del clúster [4Fe4S] en el heterodímero de la primasa de ADN intacto.
- Para entender cómo el ADN y la unión NTP influyen en la señalización electroquímica del grupo.
- Explorar la relación entre las propiedades redox y la actividad catalítica de la primasa.
Principales métodos:
- Se empleó la electroquímica del ADN para estudiar el heterodímero completo de la primasa p48/p58.
- Se realizaron mediciones electroquímicas para analizar el potencial redox del grupo [4Fe4S].
- Se realizaron ensayos de actividad enzimática para evaluar el impacto de la unión de NTP en la catálisis.
Principales resultados:
- La robusta señalización redox del grupo [4Fe4S] requería la unión simultánea de ADN y NTP.
- La unión de NTP cambió el potencial redox del grupo a un rango fisiológico (~ 160 mV vs NHE).
- La precarga de primasa con NTPs aumentó la actividad catalítica, lo que indica una conformación más activa.
Conclusiones:
- La unión al sustrato (ADN y NTP) y los cambios conformacionales regulan la señalización redox del racimo [4Fe4S].
- Las interacciones electrostáticas y la alineación de las subunidades influyen en el entorno y la señalización del grupo.
- Estos hallazgos son relevantes para la comprensión de la señalización en enzimas de procesamiento de ADN de múltiples subunidades con grupos [4Fe4S].
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