Binding Site Geometry and Subdomain Valency Control Effects of Neutralizing Lectins on HIV‑1 Viral Particles

Carbohydrate binding proteins such as griffithsin, cyanovirin-N, and BanLec are potent HIV entry inhibitors and promising microbicides. Each binds to high-mannose glycans on the surface envelope glycoprotein gp120, yet the mechanisms by which they engage viral spikes and exhibit inhibition constants ranging from nanomolar to picomolar are not understood. To determine the structural and mechanistic basis for recognition and potency, we selected a panel of lectins possessing different valencies per subunit, oligomeric states, and relative orientations of carbohydrate binding sites to systematically probe their contributions to inhibiting viral entry. Cryo-electron micrographs and immuno gold staining of lectin-treated viral particles revealed two distinct effectsnamely, viral aggregation or clustering of the HIV-1 envelope on the viral membranethat were dictated by carbohydrate binding site geometry and valency. “Sandwich” surface plasmon resonance experiments revealed that a second binding event occurs only for those lectins that could aggregate viral particles. Furthermore, picomolar <i>K</i>_d values were observed for the second binding event, providing a mechanism by which picomolar IC₅₀ values are achieved. We suggest that these binding and aggregation phenomena translate to neutralization potency