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Efectos de isótopos de átomos pesados en la reacción catalizada por el oxalato descarboxilasa de Bacillus subtilis
Laurie A Reinhardt1, Drazenka Svedruzic, Christopher H Chang
1Department of Chemistry, University of Florida, Gainesville, Florida 32611-7200, USA.
Journal of the American Chemical Society
|January 30, 2003
Resumen
La descarboxilasa de oxalato (OxDC) utiliza manganeso y dioxígeno para dividir el oxalato, produciendo formato y CO2. Los efectos cinéticos de los isótopos sugieren un mecanismo radical que involucra oxalato y formatos intermedios.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Enzimología Enzimología.
- La cinética química es la cinética química.
Sus antecedentes:
- La descarboxilasa de oxalato (OxDC) cataliza la escisión del oxalato en dióxido de carbono y formato.
- La enzima de Aspergillus niger y Bacillus subtilis contiene Mn (II) y requiere dioxígeno.
- Se ha propuesto un mecanismo de radicales libres, pero no se ha probado definitivamente.
Objetivo del estudio:
- Para investigar el mecanismo catalítico del OxDC bacteriano utilizando efectos de isótopos cinéticos de átomos pesados.
- Proporcionar evidencia experimental a favor o en contra de una vía radical intermedia.
Principales métodos:
- Las mediciones del efecto del isótopo cinético del átomo pesado se realizaron en el recombinante Bacillus subtilis OxDC.
- Los oxalatos etiquetados isotópicamente se utilizaron para medir los efectos de los isótopos (13)C y (18)O en diferentes valores de pH.
- Análisis de los efectos de isótopos normales e inversos para dilucidar los pasos de reacción.
Principales resultados:
- Se observaron efectos normales del isótopo (13) C tanto para la producción de formato como de CO2 a pH 4.2 y 5.7.
- Se observaron efectos normales (18) de isótopos O para la formación de formato en ambos valores de pH.
- Se observó un efecto isotópico inverso (18) O para la formación de CO2 a pH 5.7.
Conclusiones:
- Los efectos de isótopos cinéticos observados apoyan un mecanismo de varios pasos que involucra radicales intermedios.
- Una transferencia de electrones acoplados a protones genera un radical oxalato, seguido de una descarboxilación a un radical formato.
- La posterior reducción y protonación producen el producto final en formato.
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