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La multiplicidad de las conformaciones de las subunidades reguladoras define el conjunto estructural de la proteína

Varun Venkatakrishnan¹, Tatiana N Laremore², Theresa S C Buckley¹

¹Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.

Journal of the American Chemical Society

|April 17, 2025

Resumen

La proteína quinasa A (PKA) se restablece formando una holoenzima, en la que participan las fosfodiesterasas (PDEs) y las subunidades reguladoras. Este proceso revela múltiples conformaciones de la holoenzima PKA de reinicio.

Área de la Ciencia:

La bioquímica
Biología molecular
Señalización celular

Sus antecedentes:

La proteína quinasa A (PKA) es una enzima crucial regulada por el monofosfato cíclico de adenosina (cAMP).
El mecanismo por el cual el PKA vuelve a su estado inactivo después de la activación mediada por cAMP no se comprende completamente.
Comprender la terminación de la señal PKA es vital para comprender la regulación celular.

Objetivo del estudio:

Para aclarar el mecanismo de terminación y reinicio de la señal PKA de tipo I.
Caracterizar el conjunto estructural de la holoenzima PKA de reinicio.
Investigar el papel de las fosfodiesterasas (PDEs) en la regulación de la PKA.

Principales métodos:

Se empleó la microscopía electrónica integrativa (cryo-EM) para determinar las estructuras.
Se utilizó la espectrometría de masas estructural para analizar complejos de holoenzimas.
Se realizaron ensayos bioquímicos para estudiar la cinética y las interacciones de las enzimas.

Principales resultados:

Se identificó un mecanismo asistido por la subunidad catalítica (Cα) para el restablecimiento de la PKA de tipo I.
Se resolvieron múltiples conformaciones distintas de la holoenzima PKA libre de cAMP (RIα2:Cα2).
Se caracterizó la dinámica de la subunidad reguladora (RIα), incluido su enlace N-terminal y el dominio CNB-B.
Se destacó la interacción entre RIα, Cα y PDE8 en los señalosomas.

Conclusiones:

Las fosfodiesterasas (PDEs) juegan un papel esencial en la terminación de la señalización cAMP-PKA al facilitar el restablecimiento de la holoenzima.
La holoenzima de reinicio PKA existe como un conjunto de conformaciones impulsadas por la dinámica RIα.
Este estudio proporciona un nuevo paradigma para la regulación mediada por PDE de las vías de señalización cAMP-PKA.

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