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Polymer Microarrays for High Throughput Discovery of Biomaterials
Published on: January 25, 2012
Substrate arrays for fluorescence-based enzyme fingerprinting and high-throughput screening.
1Department of Chemistry and Biochemistry, University of Berne, Freiestrasse 3, 3012 Berne, Switzerland. jean-louis.reymond@ioc.unibe.ch
Annals of the New York Academy of Sciences
|December 22, 2007
Summary
Researchers developed new fluorogenic substrates for enzyme activity screening. These probes enable high-throughput analysis of various enzymes, aiding in drug discovery and biocatalysis development.
Area of Science:
- Biochemistry and Chemical Biology
- Enzyme Engineering and Directed Evolution
- Synthetic Chemistry and Probe Development
Background:
- Enzyme activity screening is crucial for biocatalysis and drug discovery.
- Existing methods often lack efficiency, specificity, or throughput.
- Fluorogenic substrates offer a sensitive detection method for enzyme activity.
Purpose of the Study:
- To develop novel fluorogenic substrates for a wide range of enzymes.
- To enable simultaneous, parallel screening of enzyme activities using substrate arrays.
- To facilitate high-throughput screening (HTS) for enzyme discovery and engineering.
Main Methods:
- Utilized indirect release of fluorogenic phenols (e.g., umbelliferone) as a core chemical principle.
- Synthesized diverse fluorogenic probes by linking enzyme-reactive groups to fluorescent reporters.
- Employed parallel formats like microtiter plates and substrate cocktails for simultaneous assays.
- Incorporated indirect product sensors for non-labeled substrates, including assays for diols and amino acids.
Main Results:
- Generated fluorescent probes for enzymes involved in asymmetric synthesis, including aldolase catalytic antibodies, transaldolases, alcohol dehydrogenases, lipases, epoxide hydrolases, proteases, and Bayer-Villiger monoxygenases.
- Demonstrated the ability to record enzyme-specific activity patterns using arrays of structurally related substrates.
- Showcased the utility of indirect product sensors for assaying non-labeled substrates.
- Many substrates and sensors are readily available or synthesized in few steps.
Conclusions:
- The developed fluorogenic substrates and sensors provide a versatile platform for enzyme activity profiling.
- This approach significantly enhances the efficiency and scope of high-throughput enzyme screening.
- The methodology supports enzyme discovery, engineering, and applications in biocatalysis and drug development.
