Expression and characterization of a single-chain polypeptide analogue of the human immunodeficiency virus type 1 gp120-CD4 receptor complex

The infection of CD4(+) host cells by human immunodeficiency virus type 1 ( HIV-1) is initiated by a temporal progression of interactions between speci fic cell surface receptors and the viral envelope protein, gp120. These int eractions produce a number of intermediate structures with distinct conform ational, functional, and antigenic features that may provide important targ ets for therapeutic and vaccination strategies against HIV infection. One s uch intermediate, the gp120-CD4 complex, arises from the interaction of gp1 20 with the CD4 receptor and enables interactions with specific coreceptors needed for viral entry. gp120-CD4 complexes are thus promising targets for anti-HIV vaccines and therapies. The development of such strategies would be greatly facilitated by a means to produce the gp120-CD4 complexes in a w ide variety of contexts. Accordingly, we have developed single-chain polype ptide analogues that accurately replicate structural, functional, and antig enic features of the gp120-CD4 complex. One analogue (FLSC) consists of ful l-length HIV-1BaL gp120 and the D1D2 domains of CD4 joined by a 20-amino-ac id linker. The second analogue (TcSC) contains a truncated form of the gp12 0 lacking portions of the Cl, C5, Vl, and V2 domains. Both molecules exhibi ted increased exposure of epitopes in the gp120 coreceptor-binding site but did not present epitopes of either gp120 or CD4 responsible for complex fo rmation. Further, the FLSC and TcSC analogues bound specifically to CCR5 (R 5) and blocked R5 virus infection. Thus, these single-chain chimeric molecu les represent the first generation of soluble recombinant proteins that mim ic the gp120-CD4 complex intermediate that arises during HIV replication.