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Proceedings Paper

Directed evolution of enzymes using microfluidic chips
Author(s): Zdeněk Pilát; Jan Ježek; Filip Šmatlo; Jan Kaňka; Pavel Zemánek
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Paper Abstract

Enzymes are highly versatile and ubiquitous biological catalysts. They can greatly accelerate large variety of reactions, while ensuring appropriate catalytic activity and high selectivity. These properties make enzymes attractive biocatalysts for a wide range of industrial and biomedical applications. Over the last two decades, directed evolution of enzymes has transformed the field of protein engineering. We have devised microfluidic systems for directed evolution of haloalkane dehalogenases in emulsion droplets. In such a device, individual bacterial cells producing mutated variants of the same enzyme are encapsulated in microdroplets and supplied with a substrate. The conversion of a substrate by the enzyme produced by a single bacterium changes the pH in the droplet which is signalized by pH dependent fluorescence probe. The droplets with the highest enzymatic activity can be separated directly on the chip by dielectrophoresis and the resultant cell lineage can be used for enzyme production or for further rounds of directed evolution. This platform is applicable for fast screening of large libraries in directed evolution experiments requiring mutagenesis at multiple sites of a protein structure.

Paper Details

Date Published: 23 December 2016
PDF: 7 pages
Proc. SPIE 10142, 20th Slovak-Czech-Polish Optical Conference on Wave and Quantum Aspects of Contemporary Optics, 101420A (23 December 2016); doi: 10.1117/12.2264377
Show Author Affiliations
Zdeněk Pilát, Institute of Scientific Instruments of the CAS, v.v.i. (Czech Republic)
Jan Ježek, Institute of Scientific Instruments of the CAS, v.v.i. (Czech Republic)
Filip Šmatlo, Institute of Scientific Instruments of the CAS, v.v.i. (Czech Republic)
Jan Kaňka, Institute of Scientific Instruments of the CAS, v.v.i. (Czech Republic)
Pavel Zemánek, Institute of Scientific Instruments of the CAS, v.v.i. (Czech Republic)


Published in SPIE Proceedings Vol. 10142:
20th Slovak-Czech-Polish Optical Conference on Wave and Quantum Aspects of Contemporary Optics
Jarmila Müllerová; Dagmar Senderáková; Libor Ladányi; Ľubomír Scholtz, Editor(s)

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