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Efficient Purification and LC-MS/MS-based Assay Development for Ten-Eleven Translocation-2 5-Methylcytosine Dioxygenase
Published on: October 15, 2018
Una enzima faltante en la biosíntesis de tiamina tiazol: identificación de TenI como una tiazol tautomerasa
Amrita B Hazra1, Ying Han, Abhishek Chatterjee
1Department of Chemistry, Texas A&M University, College Station, Texas 77843, USA.
Journal of the American Chemical Society
|May 4, 2011
Resumen
La enzima bacteriana TenI, de función previamente desconocida, es identificada como una tiazol tautomerasa. Este descubrimiento completa el conjunto de enzimas requerido para la principal vía de biosíntesis de tiamina bacteriana.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Biología Molecular Biología Molecular
- Enzimología Enzimología.
Sus antecedentes:
- La enzima TenI se encuentra frecuentemente con genes para la biosíntesis de tiamina tiazol en bacterias.
- A pesar de la similitud de la secuencia con la tiamina fosfato sintasa, TenI carece de esta actividad, dejando su papel en la producción de tiamina poco clara.
Objetivo del estudio:
- Para aclarar la función desconocida de la enzima bacteriana TenI.
- Caracterizar el mecanismo catalítico y la estructura de TenI en relación con la biosíntesis de tiamina.
Principales métodos:
- Pruebas de purificación de enzimas y de actividad.
- Cristalografía de rayos X para determinar la estructura del complejo enzima-ligando.
- Mutagénesis dirigida al sitio para identificar residuos catalíticos.
Principales resultados:
- TenI funciona como una tiazol tautomerasa, no como una tiamina fosfato sintasa.
- La estructura del complejo enzima-sustrato reveló el fosfato y la histidina 122 como componentes catalíticos clave.
- Se propuso un mecanismo de reacción que involucra a la histidina 122 como catalizador ácido/base.
Conclusiones:
- La función de TenI como una tiazol tautomerasa ahora está establecida.
- Este hallazgo resuelve la identidad de todas las enzimas en la vía de biosíntesis de tiamina bacteriana primaria.
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