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E. coli surface display of streptavidin for directed evolution of an allylic deallylase

Bernardes Pinheiro, VB; (2018) E. coli surface display of streptavidin for directed evolution of an allylic deallylase. Chemical Science , 9 (24) pp. 5383-5388. 10.1039/C8SC00484F. Green open access

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Abstract

Artificial metalloenzymes (ArMs hereafter) combine attractive features of both homogeneous catalysts and enzymes and offer the potential to implement new-to-nature reactions in living organisms. Herein we present an E. coli surface display platform for streptavidin (Sav hereafter) relying on an Lpp-OmpA anchor. The system was used for the high throughput screening of a bioorthogonal CpRu-based artificial deallylase (ADAse) that uncages an allylcarbamate-protected aminocoumarin 1. Two rounds of directed evolution afforded the double mutant S112M–K121A that displayed a 36-fold increase in surface activity vs. cellular background and a 5.7-fold increased in vitro activity compared to the wild type enzyme. The crystal structure of the best ADAse reveals the importance of mutation S112M to stabilize the cofactor conformation inside the protein.

Type: Article
Title: E. coli surface display of streptavidin for directed evolution of an allylic deallylase
Open access status: An open access version is available from UCL Discovery
DOI: 10.1039/C8SC00484F
Publisher version: http://doi.org/10.1039/C8SC00484F
Language: English
Additional information: Copyright This journal is © The Royal Society of Chemistry 2018. Open Access Article. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. - With correction dated 28 June 2019.
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Structural and Molecular Biology
URI: https://discovery.ucl.ac.uk/id/eprint/10049875
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