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Superficies de interacción de proteínas localizadas en la proteína portadora de EntB reveladas por mutagénesis
Jonathan R Lai1, Michael A Fischbach, David R Liu
1Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Journal of the American Chemical Society
|August 24, 2006
Resumen
Las proteínas portadoras son clave para el ensamblaje de la poliquetida sintasa (PKS) y la peptídica sintetasa no ribosómica (NRPS). Las superficies de interacción localizadas en EntB-ArCP facilitan la unión de la fosfopanteteinil transferasa, distinta de otras interacciones del módulo.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Biología Molecular Biología Molecular
- Enzimología Enzimología.
Sus antecedentes:
- Las proteínas portadoras son componentes esenciales de las líneas de ensamblaje de la policetida sintasa (PKS) y la peptídica sintetasa no ribosómica (NRPS).
- Estas proteínas presentan intermediarios biosintéticos como tioesters, que requieren interacciones específicas proteína-proteína para una catálisis eficiente.
- Comprender estas interacciones es crucial para la ingeniería de nuevos sistemas PKS y NRPS.
Objetivo del estudio:
- Para identificar las superficies de interacción de proteínas localizadas en la proteína portadora de arilo de EntB (EntB-ArCP).
- Investigar el papel de los residuos específicos en la mediación de las interacciones con las fosfopanteteiniltransferasas (PPTasas).
- Explorar el potencial para diseñar interacciones proteína-proteína alteradas en PKS y NRPS.
Principales métodos:
- Mutagénesis combinatoria de las regiones N-terminales de EntB-ArCP.
- Selección para la producción funcional de enterobactinas en medios con deficiencia de hierro.
- Identificación de residuos conservados involucrados en las interacciones de las proteínas.
Principales resultados:
- Se identificaron superficies de interacción de proteínas localizadas en EntB-ArCP.
- Se encontró que los residuos G242 y D244 forman una superficie de interacción para las PPTases EntD y Sfp.
- Esta interfaz PPTase es distinta de la interfaz previamente conocida para el módulo de alargamiento EntF.
- La mayoría de los residuos superficiales de EntB-ArCP no están involucrados en estas interacciones caracterizadas.
Conclusiones:
- EntB-ArCP posee distintas superficies de interacción para diferentes socios de proteínas.
- Los componentes de las proteínas reconocen diferentes caras de EntB-ArCP dentro de la enterobactina sintetasa.
- El diseño de interacciones de proteínas portadoras no cognadas en los sistemas PKS y NRPS es factible con mutaciones mínimas.
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