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

Heinisch, Tillmann and Schwizer, Fabian and Garabedian, Brett and Csibra, Eszter and Jeschek, Markus and Vallapurackal, Jaicy and Pinheiro, Vitor B. and Marlière, Philippe and Panke, Sven and Ward, Thomas R.. (2018) E. coli surface display of streptavidin for directed evolution of an allylic deallylase. Chemical Science, 9 (24). pp. 5383-5388.

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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.
Faculties and Departments:05 Faculty of Science > Departement Chemie > Chemie > Bioanorganische Chemie (Ward)
UniBasel Contributors:Ward, Thomas R. R. and Heinisch, Tillmann and Schwizer, Fabian and Garabedian, Brett and Vallapurackal, Jaicy and Panke, Sven
Item Type:Article, refereed
Article Subtype:Research Article
Publisher:Royal Society of Chemistry
ISSN:2041-6520
e-ISSN:2041-6539
Note:Publication type according to Uni Basel Research Database: Journal article
Language:English
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Last Modified:23 May 2019 14:06
Deposited On:22 May 2019 12:00

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