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Identificación del entorno de valencia y coordinación de los centros de cobre de las partículas metano monooxigenasa
Megen A Culpepper1, George E Cutsail, William A Gunderson
1Departments of ‡Molecular Biosciences and of §Chemistry, Northwestern University , Evanston, Illinois 60208, United States.
Journal of the American Chemical Society
|July 26, 2014
Resumen
La monooxigenasa de metano en partículas (pMMO) utiliza centros de cobre para oxidar el metano. Este estudio identifica un ion de cobre EPR-activo en el sitio de dicopio, aclarando pMMO.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Enzimología Enzimología.
- La espectroscopia es una técnica de espectroscopia.
Sus antecedentes:
- La monooxigenasa de metano en partículas (pMMO) es crucial para la oxidación del metano en las bacterias metanotróficas.
- Sus centros de cobre son clave para su funcionamiento, pero son difíciles de caracterizar utilizando la espectroscopia EPR debido a los múltiples sitios de cobre.
Objetivo del estudio:
- Para aclarar las funciones y los entornos de coordinación de los centros de cobre en pMMO utilizando métodos espectroscópicos.
- Validar un fragmento recombinante soluble (spmoB) como un modelo confiable para el estudio de pMMO.
Principales métodos:
- La espectroscopia de resonancia paramagnética de electrones (EPR, por sus siglas en inglés) es una espectroscopia de resonancia paramagnética de electrones.
- La espectroscopia de doble resonancia nuclear de electrones (ENDOR) es una espectroscopia de doble resonancia nuclear de electrones.
- Modulación de la envoltura de eco de espín de electrones (ESEEM) espectroscopia espectroscopia.
- Análisis del contenido de metales.
- Mutagenesis dirigida al sitio de las variantes de spmoB.
Principales resultados:
- Los datos espectroscópicos de spmoB coincidieron estrechamente con el pMMO nativo, lo que confirma que spmoB es un modelo válido.
- Se identificó un ion EPR-activo de Cu (II) por pMMO, ubicado dentro del sitio activo del dicoplo como un par de Cu (I) localizado en la valencia.
- El sitio de monocopio fue asignado como Cu(I).
- Residuos específicos de histidina (His137/His139 o His33/N-terminal) estuvieron implicados en la coordinación de Cu.
- No se encontraron ligandos acuáticos unidos al ion Cu (II).
Conclusiones:
- El estudio aclara las identidades de los sitios de cobre y la coordinación en pMMO.
- Proporciona un sistema de modelo validado (spmoB) para futuras investigaciones de los mecanismos de oxidación del metano.
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