Characterizing the Binding Kinetics of X31 Influenza Virus to Glycolipids using Single-Particle Tracking

Abstract

The infection process for enveloped viruses begins with the attachment of viruses to the host cell surface, followed by the fusion of the viral membrane with the host membrane. For Influenza, attachment and fusion are mediated by a single protein, hemagglutinin (HA), anchored in the viral membrane. Successful virus attachment to the host cell hinges on several factors such as the chemical affinity of HA to sialic acid (SA) receptors, polyvalent binding, and binding site accessibility. To characterize binding kinetics of influenza X31 (H3N2), total internal reflection microscopy is used to observe dye-labeled viruses interacting with SA receptors embedded in a supported lipid bilayer. An important advantage of the single-particle binding assay is that it can extract the binding and unbinding times of individual viruses, which are needed to determine the binding residence time distribution and binding rate constant (kon). The binding residence time distribution is used to characterize the binding affinity of HA to SA and binding avidity of a virus to several SA receptors. Meanwhile, the binding rate constant is used to quantify the accessibility of HA to SA.In this work, we examine influenza binding to several types of glycolipids. The glycolipids are GM3, GM1, and GD1a, which all have the alpha 2-3 linked sialic acid group but with varying peripheral glycan structures. Results show that the binding rate is the greatest for GD1a, followed by GM3 and then GM1. The binding residence time data, however, shows little difference between the three receptors. We will discuss possible rationales for these observations, while demonstrating the utility of the single particle assay as a convenient tool for studying binding kinetics.