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Multi-enzyme Screening Using a High-throughput Genetic Enzyme Screening System

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High-throughput metabolic engineering: advances in small-molecule screening and selection.

Jeffrey A Dietrich¹, Adrienne E McKee, Jay D Keasling

¹UCSF-UCB Joint Graduate Group in Bioengineering, Berkeley, California 94720, USA. jadietrich@berkeley.edu

Annual Review of Biochemistry

|April 7, 2010

Summary

Directed evolution requires high-throughput screening for small molecule overproduction. This review details colorimetric, fluorescent, and biosensor methods to detect hard-to-find molecules, enhancing metabolic engineering efforts.

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Area of Science:

Metabolic Engineering
Synthetic Biology
Biotechnology

Background:

Metabolic engineering aims to overproduce valuable small molecules, but many are difficult to screen for.
Directed evolution is crucial for improving production titers but is limited by screening capabilities.

Purpose of the Study:

To review high-throughput screening techniques for detecting inconspicuous small molecules.
To establish a screening and selection ladder based on assay throughput and sensitivity.
To analyze current and future small-molecule detection methods in directed evolution.

Main Methods:

Analysis of colorimetric, fluorescent, and growth-coupled screening techniques.
Evaluation of in vivo biosensor-based detection methods.
Discussion of constraints in standard directed evolution workflows.

Main Results:

High-throughput screening methods enable the detection of previously inaccessible small molecules.
In vivo biosensors reduce in vitro manipulations and increase screening throughput.
A screening and selection ladder optimizes throughput and sensitivity.

Conclusions:

Development of advanced screening techniques is vital for metabolic engineering.
Biosensor-based and transcription factor-based systems offer promising avenues for future small-molecule detection.
These advancements facilitate the overproduction of high-value compounds.