Direct visualization of the oligomeric state of hemagglutinins of influenza virus by high-resolution atomic force microscopy.

High-resolution atomic force microscopy (AFM) is a powerful technique for the direct visualization of single molecules. Here, AFM is applied to characterize the oligomeric state of hemagglutinins of the influenza virus. Hemagglutinins are known to be present in a trimeric form inside the viral envelope. However, recombinant hemagglutinins are also present as large oligomers, which impair the functional activity of the protein. Five commercial recombinant hemagglutinins from the viral strains H1, H3, H5, H7, and H9 were studied with high-resolution AFM. Functionally inactive hemagglutinins were shown to have a higher percentage of large oligomers compared with the proteins with functional activity. Large oligomers were revealed to be unstable; the oligomeric state of hemagglutinin was affected by pH or the presence of ligands. Antibody binding shifts the balance to small oligomers, whereas DNA aptamer induced the formation of large associates.