Synthetic RNA switches for yeast metabolic engineering: Screening recombinant enzyme libraries

Joshua K. Michener, Christina D. Smolke

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

Directed evolution is a powerful technique for increasing the activity of poorly active enzymes, for example when an enzyme is engineered to accept a new substrate or function in a new environment. Since enzyme activity greatly depends on the enzyme environment, screening should be performed under the same conditions as the ultimate application of the enzyme. When an enzyme will be used in live cells, RNA biosensors offer a powerful and fl exible method of linking the desired phenotype, production of a small molecule of interest, to an easily measured phenotype, such as fl uorescence. Here, we describe methods for screening enzyme libraries using an RNA biosensor, showing examples from the evolution of a P450 monooxygenase.

Original languageEnglish
Pages (from-to)125-136
Number of pages12
JournalMethods in molecular biology (Clifton, N.J.)
Volume1152
DOIs
StatePublished - 2014

Funding

FundersFunder number
National Center for Complementary and Integrative HealthDP1AT007886

    Keywords

    • Directed evolution
    • High-throughput enzyme screening
    • P450
    • RNA switch

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