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Los interruptores de la enzima RecBCD dirigen las subunidades del motor en respuesta al reconocimiento del chi

Maria Spies¹, Ichiro Amitani, Ronald J Baskin

¹Section of Microbiology, University of California, Davis, CA 95616-8665, USA.

|November 21, 2007

Resumen

La enzima RecBCD utiliza dos motores, RecB y RecD, para moverse a lo largo del ADN. Después de encontrarse con un

Área de la Ciencia:

Biología Molecular Biología Molecular
La bioquímica es la bioquímica.
Genética La genética.

Sus antecedentes:

RecBCD es una enzima crucial de la ADN helicasa involucrada en la reparación y recombinación del ADN.
La actividad de la enzima está regulada por el punto de acceso chi, que afecta a su velocidad de translocación.
Comprender la dinámica motora de RecBCD es clave para comprender las vías de procesamiento del ADN.

Objetivo del estudio:

Investigar las distintas funciones de las subunidades motoras RecB y RecD en la translocación de RecBCD.
Para aclarar el mecanismo por el cual el reconocimiento de chi altera el movimiento de RecBCD.
Para comparar la dinámica de translocación de RecBCD de tipo salvaje con un mutante de RecD defectuoso.

Principales métodos:

Se emplearon técnicas de visualización de una sola molécula para observar la actividad de la enzima RecBCD.
Un mutante RecBCD ((K177Q), con un motor RecD defectuoso, fue diseñado y estudiado.
El tratamiento de la nucleasa P1 se utilizó para sondear las estructuras del ADN y las interacciones enzimáticas.

Principales resultados:

RecBCD ((K177Q) hizo una pausa en el chi pero mantuvo una velocidad de translocación constante, a diferencia del RecBCD de tipo salvaje.
La tasa de translocación del mutante RecBCD ((K177Q) coincidía con la tasa post-chi de la enzima de tipo salvaje, lo que sugiere que RecB lidera después de chi.
El tratamiento con nucleasa P1 reveló un bucle de ssDNA que contiene chi e indicó que RecD es el motor más rápido antes de chi.

Conclusiones:

RecBCD se somete a un interruptor de motor en el punto de acceso chi, con RecD liderando antes de chi y RecB liderando después de chi.
Este mecanismo de conmutación del motor puede coordinar los procesos de degradación y reparación del ADN.
Las diferentes velocidades de translocación son críticas para regular la carga de RecA durante la reparación del ADN.

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