Related Experiment Video
Updated: Dec 20, 2025
08:10
Multi-enzyme Screening Using a High-throughput Genetic Enzyme Screening System
Published on: August 8, 2016
9.2K
A High-Throughput Screening System Based on Droplet Microfluidics for Glucose Oxidase Gene Libraries
Radivoje Prodanović1,2, W Lloyd Ung2, Karla Ilić Đurđić1
1Faculty of Chemistry, University of Belgrade, Studentski trg 12, 11000 Belgrade, Serbia.
Molecules (Basel, Switzerland)
|May 28, 2020
Summary
Researchers developed a novel microfluidic platform for screening glucose oxidase (GOx) enzyme mutants. This ultrahigh-throughput system significantly enhances the efficiency of enzyme optimization for industrial applications.
Area of Science:
- Biotechnology
- Enzyme Engineering
- Microfluidics
Background:
- Glucose oxidase (GOx) is a key industrial enzyme.
- Mutagenesis and activity-based screening are crucial for optimizing GOx.
- Existing screening methods have limitations in efficiency and throughput.
Purpose of the Study:
- To develop an ultrahigh-throughput screening platform for GOx mutants.
- To improve the efficiency of GOx optimization using a microfluidic lab-on-chip device.
- To identify novel GOx mutants with enhanced activity.
Main Methods:
- Utilized a microfluidic lab-on-chip device for ultrahigh-throughput screening.
- Expressed GOx mutants on the surface of yeast cells.
- Measured GOx activity via fluorescence detection in microdroplets using coupled enzyme reactions.
- Employed a saturation mutagenesis library for GOx variants.
Main Results:
- Achieved a >35-fold enrichment of GOx mutants with higher than wild-type activity in two sorting rounds.
- Demonstrated a nearly twofold increase in efficiency compared to previous flow cytometry platforms.
- Identified novel GOx mutants, including M6 with six point mutations.
- The M6 mutant exhibited a 2.1-fold increase in kcat compared to wild-type GOx.
Conclusions:
- The new microfluidic platform is more sensitive and efficient than flow cytometry for GOx screening.
- This platform enables comprehensive screening of gene libraries with multiple mutations.
- The developed platform accelerates the discovery of improved industrial enzymes like GOx.
