Further evidence for hexagonal organization of HIV gag protein in prebudding assemblies and immature virus-like particles.

The fullerene-like model for the organization of HIV gag encoded precursor pr55gag was based on the study of prebudding assemblies at the plasma membrane of cells infected with a recombinant baculovirus expressing HIV-1 gag protein. The objective of the present study was to support the model by image processing of virus-like particles (VLP). In this work we used VLP purified by density gradient centrifugation, which caused partial or occasionally complete loss of the lipid bilayer in some VLP without the use of detergent. In addition more plasma membrane-associated pr55gag protein assemblies were processed. Image processing of negatively stained specimens revealed the presence of threefold symmetry and a hexagonal network of rings with a resolution of 29 A in VLP and better than 25 A in membrane associated assemblies. The center-to-center spacing of the rings was 67 A in VLP and 70 A in membrane assemblies. Patches of gag protein oligomers at the plasma membrane were usually round and varying in size, but some of them were triangular. Indication of triangular-shaped gag protein assemblies was also seen in partly dissociated VLP. Since the hexagonal network is formed by the uncleaved gag polyprotein, we conclude that the threefold symmetry applies to all domains including p24gag. The presence of threefold symmetry and the hexagonal network in VLP are consistent with the hypothesis that immature HIV particles possess icosahedral symmetry.