We have constructed DNA vectors expressing two soluble forms of the SF162 envelope. Both forms comprise the entire gp120 subunit and the extracellular part of the gp41 subunit. In the one construct the gp120-gp41 cleavage site was mutated so that only one polypeptide of 140kd is produced ('fused' protein). In the other construct the gp120-gp41 cleavage site remained intact so two polypeptides (120kD and 20kD) are produced ('cleaved' protein). Upon transient transfection of 193T cells, both the 'fused' and 'cleaved' proteins are secreted in the cell culture medium as oligomers. Both proteins are precipitated by HIV+ human sera and several gp120 and gp41 antibodies that bind to linear and conformation-dependent epitopes. In addition, the relative CD4-binding affinity of these two proteins is lower that that of monomeric gp120 and similar to that of intact SF162 virions. These preliminary studies suggest therefore that the overall conformation of these two proteins resembles more that found on infectious SF162 virions than the corresponding monomeric gp120. We are currently examining the type and extent of conformational changes undergone by these two proteins upon CD4 binding and comparing these changes to those occurring on the surface of SF162 virions. These proteins might be useful as immunogens to elicit antibody responses against SF162 and other primary isolates.
Keywords: AEGIS, HIV-1, Antigens, CD4, Virion, Epitopes, Human, analysis, AIDS
