The envelope protein is an important determinant of HIV-1 cell-specific tropism. The gp160 envelope precursor proteins from macrophage-tropic or T lymphoid cell line-tropic strains of HIV-1 were expressed in recombinant vaccinia virus-infected cell lines or primary lymphocytes or macrophages. No significant differences in the kinetics of synthesis of gp160, processing into gp120 and gp41 proteins, N-linked glycosylation, or release of gp120 into the medium were noted with the different envelope proteins. However, gp120 envelope protein shed into the medium was found to be at least partially cleaved at a site within the V3 loop. The gp120 envelope proteins from macrophage-tropic isolates exhibited lower rates of cleavage than those from lymphoid cell line-tropic strains in all cell types examined. Cell-free protease digestion studies also demonstrated relative resistance of the envelopes from macrophage-tropic compared to lymphoid cell line-tropic strains. All recombinant envelope proteins were recognized by monoclonal antibodies directed at gp41 or the C-terminal gp120 epitopes, and no differences in binding to CD4 were noted.