A Robust Growth-Based Selection Platform to Evolve an Enzyme via Dependency on Noncanonical Tyrosine Analogues

Suzanne C. Jansen, Clemens Mayer*

*Bijbehorende auteur voor dit werk

OnderzoeksoutputAcademicpeer review

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Samenvatting

Growth-based selections evaluate the fitness of individual organisms at a population level. In enzyme engineering, such growth selections allow for the rapid and straightforward identification of highly efficient biocatalysts from extensive libraries. However, selection-based improvement of (synthetically useful) biocatalysts is challenging, as they require highly dependable strategies that artificially link their activities to host survival. Here, we showcase a robust and scalable growth-based selection platform centered around the complementation of noncanonical amino acid-dependent bacteria. Specifically, we demonstrate how serial passaging of populations featuring millions of carbamoylase variants autonomously selects biocatalysts with up to 90,000-fold higher initial rates. Notably, selection of replicate populations enriched diverse biocatalysts, which feature distinct amino acid motifs that drastically boost carbamoylase activity. As beneficial substitutions also originated from unintended copying errors during library preparation or cell division, we anticipate that our growth-based selection platform will be applicable to the continuous, autonomous evolution of diverse biocatalysts in the future.

Originele taal-2English
Artikelnummer4c00070
Pagina's (van-tot)1583–1590
Aantal pagina's8
TijdschriftJACS Au
Volume4
Nummer van het tijdschrift4
Vroegere onlinedatum19-mrt.-2024
DOI's
StatusPublished - 22-apr.-2024

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