A universal flow cytometry assay for screening carbohydrate-active enzymes using glycan microspheres

Area of Science:

• Biochemistry
• Enzymology
• Glycobiology

Background:

• Carbohydrate-active enzymes (CAZymes) play crucial roles in biological processes.
• Screening CAZyme activities is essential for understanding their functions and applications.
• Existing methods for CAZyme activity screening can be complex and time-consuming.

Purpose of the Study:

• To develop a simple, multiplexed assay for screening carbohydrate-active enzyme activities.
• To integrate multiple steps including glycan synthesis, bioconjugation, detection, and analysis into a single workflow.
• To enable high-throughput screening of diverse CAZyme families.

Main Methods:

• The assay integrates glycan synthesis and bioconjugation to microspheres.
• Fluorescent chemical and biochemical detection methods are employed.
• Multiparameter flow cytometric analysis is used for data acquisition and interpretation.

Main Results:

• The developed assay successfully integrates multiple steps for efficient CAZyme activity screening.
• The multiplexed format allows for simultaneous analysis of different enzyme families.
• Flow cytometry provides sensitive and quantitative detection of enzyme activities.

Conclusions:

• This assay offers a simple and efficient platform for screening carbohydrate-active enzyme activities.
• The integrated approach streamlines the process, saving time and resources.
• The method is adaptable for studying various CAZymes and their functions.