Engineered OAA lectins as selective and sensitive high mannose glycan targeting tools

Area of Science:

• Carbohydrate chemistry
• Protein engineering
• Structural biology

Background:

• Oscillatoria agardhii agglutinin (OAA) lectin binds to high mannose N-glycans (HMGs) via a shared pentamannose core.
• Existing tools lack the sensitivity to differentiate specific HMG structures within their biological context due to subtle structural variations.

Purpose of the Study:

• To engineer OAA variants with enhanced selectivity for specific HMG structures, particularly Man ₅ GlcNAc ₂.
• To develop a structure-specific detection tool for HMGs.
• To explore OAA variants as potential antiviral agents.

Main Methods:

• Phage display screening of a diverse OAA variant library.
• Site-directed mutagenesis and binding analysis of OAA variants.
• Crystal structure determination of a selective OAA variant.
• Valency modulation of engineered OAA variants on a beta-barrel scaffold.

Main Results:

• Identified an OAA variant with high selectivity for Man ₅ GlcNAc ₂.
• Discovered four key mutations conferring selectivity and two enhancing overall HMG affinity.
• Elucidated the mechanism of co-dependent mutations in achieving selectivity through structural and binding analyses.
• Demonstrated significant improvement in binding properties through valency modulation.
• Showcased engineered OAA variants as effective HMG profiling tools and tunable antiviral agents.

Conclusions:

• Engineered OAA variants offer precise detection of specific HMG structures.
• The study provides insights into mutation-driven selectivity and affinity enhancement in lectins.
• Developed OAA variants hold promise for improved glycan analysis and antiviral therapies.