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A Practical Guide to Phage- and Robotics-Assisted Near-Continuous Evolution

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Un sistema para la evolución dirigida continua de las biomoléculas.

Kevin M Esvelt¹, Jacob C Carlson, David R Liu

¹Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138, USA.

|April 12, 2011

Resumen

La evolución continua asistida por fagos (PACE, por sus siglas en inglés) permite una rápida y continua evolución en laboratorio de moléculas codificadas por genes en bacterias. Este avance acelera el descubrimiento de nuevas biomoléculas y enzimas, mejorando significativamente la eficacia evolutiva.

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Área de la Ciencia:

Biología Molecular Biología Molecular
Biotecnología La biotecnología es la biotecnología.
Biología evolutiva Biología evolutiva.

Sus antecedentes:

La evolución de laboratorio acelera el desarrollo de biomoléculas, pero es lenta y requiere mucha mano de obra.
Los métodos anteriores de evolución continua estaban limitados a moléculas específicas como las ribozimas.
Acelerar los ciclos evolutivos es clave para mejorar la ingeniería biomolecular.

Objetivo del estudio:

Desarrollar un sistema para la evolución dirigida continua de moléculas codificadas por genes en bacterias.
Aumentar significativamente la velocidad y la eficiencia de la evolución del laboratorio.
Para permitir la ingeniería rápida de proteínas con nuevas funciones.

Principales métodos:

Sistema de evolución continua asistida por fagos (PACE) que utiliza un ciclo de vida de bacteriófago modificado en Escherichia coli.
Los genes se transfieren entre las células huésped dependiendo de la actividad molecular deseada.
PACE permite docenas de rondas de evolución por día sin intervención humana.

Principales resultados:

PACE evolucionó con éxito variantes de la RNA polimerasa (RNAP) T7 con especificidad de promotor alterada y iniciación de nucleótidos.
Las variantes de RNAP diseñadas mostraron cientos de mejoras en la actividad en comparación con las de tipo salvaje.
200 rondas de evolución se completaron en 8 días, produciendo enzimas con las actividades deseadas desde niveles indetectables.

Conclusiones:

PACE acelera dramáticamente la evolución en el laboratorio, permitiendo el rápido descubrimiento de biomoléculas funcionales.
El sistema supera las limitaciones de las técnicas anteriores de evolución dirigida.
PACE ofrece una poderosa herramienta para abordar los complejos desafíos de la ingeniería de proteínas y estudiar la evolución molecular.