Bulk Fusion Studies of Sendai Virus and Application of Mass Action Kinetic Fusion Model

Abstract

Paramyxoviruses are enveloped, negative-sense RNA viruses which include prominent pathogens such as mumps, measles, and parainfluenza viruses. For paramyxoviruses to infect a host cell, viral surface proteins must first bind to the host cell membrane and subsequently catalyze the fusion of the viral and host cell membranes to deliver the viral genome into the cell. Certain general features of the binding and fusion process are believed to be conserved within the paramyxovirus family, though it is quite diverse. There remains ambiguity in the literature concerning the fusion mechanism(s) and intermediate states. Here we study the fusion of Sendai virus as a model system. We explore the fusion of the virus with unilamellar liposomes in bulk by fluorescence spectroscopy, specifically examining the timescale of fusion and the role of the putative viral receptor, ganglioside GD1a. The virus was prepared by inserting lipophilic R-18 dye into the viral membrane and fusion with the liposomes was studied via fluorescence dequenching. The results were then analyzed using a model that is inspired by chemical mass action kinetics and which simulates the fluorescent data collected from the experiments to extract key parameters such as the rate of association between the liposomes and the virus, the number of fused liposomes per viral particle, and their rate of fusion. We observed that under our experimental conditions lipid mixing was a slow process, occurring over several hours, and we discuss the influence of the viral receptor on fusion as well as the modeling results.