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Combinatorial metabolic engineering using an orthogonal tri-functional CRISPR system.

ABSTRACT: Designing an optimal microbial cell factory often requires overexpression, knock-down, and knock-out of multiple gene targets. Unfortunately, such rewiring of cellular metabolism is often carried out sequentially and with low throughput. Here, we report a combinatorial metabolic engineering strategy based on an orthogonal tri-functional CRISPR system that combines transcriptional activation, transcriptional interference, and gene deletion (CRISPR-AID) in the yeast Saccharomyces cerevisiae. This strategy enables perturbation of the metabolic and regulatory networks in a modular, parallel, and high-throughput manner. We demonstrate the application of CRISPR-AID not only to increase the production of ?-carotene by 3-fold in a single step, but also to achieve 2.5-fold improvement in the display of an endoglucanase on the yeast surface by optimizing multiple metabolic engineering targets in a combinatorial manner.

SUBMITTER: Lian J

PROVIDER: S-EPMC5700065 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Combinatorial metabolic engineering using an orthogonal tri-functional CRISPR system.

Lian Jiazhang J HamediRad Mohammad M Hu Sumeng S Zhao Huimin H

Nature communications 20171122 1

Designing an optimal microbial cell factory often requires overexpression, knock-down, and knock-out of multiple gene targets. Unfortunately, such rewiring of cellular metabolism is often carried out sequentially and with low throughput. Here, we report a combinatorial metabolic engineering strategy based on an orthogonal tri-functional CRISPR system that combines transcriptional activation, transcriptional interference, and gene deletion (CRISPR-AID) in the yeast Saccharomyces cerevisiae. This ...[more]

PMID: 29167442

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