Widely applicable background depletion step enables transaminase evolution through solid-phase screening

Matteo Planchestainer, Eimear Hegarty, Christian M. Heckmann, Louise J. Gourlay, Francesca Paradisi*

*Corresponding author for this work

Research output: Contribution to journalArticleScientificpeer-review

16 Citations (Scopus)

Abstract

Directed evolution of transaminases is a widespread technique in the development of highly sought-after biocatalysts for industrial applications. This process, however, is challenged by the limited availability of effective high-throughput protocols to evaluate mutant libraries. Here we report a rapid, reliable, and widely applicable background depletion method for solid-phase screening of transaminase variants, which was successfully applied to a transaminase from Halomonas elongata (HEWT), evolved through rounds of random mutagenesis towards a series of diverse prochiral ketones. This approach enabled the identification of transaminase variants in viable cells with significantly improved activity towards para-substituted acetophenones (up to 60-fold), as well as tetrahydrothiophen-3-one and related substrates. Rationalisation of the mutants was assisted by determination of the high-resolution wild-type HEWT crystal structure presented herein.
Original languageEnglish
Pages (from-to)5952-5958
Number of pages7
JournalChemical Science
Volume10
Issue number23
DOIs
Publication statusPublished - 2019
Externally publishedYes

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