Glycan specificity of neuraminidases determined in microarray format

Area of Science:

• Biochemistry
• Glycobiology
• Enzymology

Background:

• Neuraminidases cleave sialic acids from glycoconjugates, playing roles in human physiology and disease.
• Understanding neuraminidase substrate specificity is crucial for elucidating their mechanisms of action.
• Current high-throughput methods for determining glycan specificity are limited.

Purpose of the Study:

• To develop and validate a high-throughput method for assessing neuraminidase activity and glycan specificity.
• To compare the efficacy of different glycan labeling reactions for monitoring neuraminidase activity.

Main Methods:

• Utilized two glycan labeling reactions: periodate oxidation and aniline-catalyzed oxime ligation (PAL) and galactose oxidase and aniline-catalyzed oxime ligation (GAL).
• Assessed neuraminidase activity against glycans in microtiter plates and on glass slide microarrays.
• Employed GAL for accurate measurement of neuraminidase activity on glycan microarrays.

Main Results:

• Both PAL and GAL could monitor neuraminidase activity in microtiter plates.
• Only GAL accurately measured neuraminidase activity on glycans immobilized on glass slide microarrays.
• GAL confirmed known specificities of three pneumococcal neuraminidases and identified novel glycan specificities.

Conclusions:

• Galactose oxidase and aniline-catalyzed oxime ligation (GAL) is a reliable method for high-throughput analysis of neuraminidase activity on glycan arrays.
• This method provides valuable insights into the glycan specificity of neuraminidases, including pneumococcal strains.
• The findings advance the understanding of neuraminidase function in biological systems.