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Lysozyme assay using a rationally designed GN4G2 substrate with coupled β-glucosidase reaction
Moe Kudo1, Haruka Kono2, Hirotake Suda1
1Graduate School of Food and Agricultural Sciences, Fukushima University, 1 Kanayagawa, Fukushima, 960-1296, Japan.
Analytical Biochemistry
|July 3, 2026
Summary
A new lysozyme assay uses a designed substrate (GN4G2) and coupled enzyme reaction for reliable measurement. This method offers improved reproducibility and accuracy for enzyme analysis.
Area of Science:
- Biochemistry
- Enzymology
Background:
- Lysozyme (EC 3.2.1.17) is crucial for host defense, cleaving peptidoglycan and chitin.
- Conventional lysozyme assays lack reproducibility due to substrate variability and assay sensitivity.
Purpose of the Study:
- Develop a simpler, more reliable lysozyme activity assay.
- Enhance reproducibility in enzyme activity measurements.
Main Methods:
- Designed a novel substrate, N-acetylchitotetraosyl cellobiose (GN4G2).
- Utilized a coupled β-glucosidase reaction for signal amplification.
- Performed molecular dynamics simulations and kinetic analysis.
Main Results:
- GN4G2 demonstrated high water solubility and catalytic efficiency.
- The assay showed linear response to time and enzyme concentration.
- Achieved good reproducibility (CV < 10%) consistent with HPLC.
Conclusions:
- The novel assay provides a robust and reproducible method for lysozyme activity determination.
- This methodology is valuable for biochemical characterization and enzyme analysis.
