Mutation in the V3 loop of HIV-1 gp120 could affect syncytium formation, virus infectivity and neutralization. To acquire more information of the V3 loop mutation, we analyzed amino acid sequences of the V3 loop of 24 504 isolates from most HIV-1 clades (including A, B, C, D, E, F, G and H clades). The consensus sequence of the V3 loop of each subtype with the highest frequency emerging on each position is constituted and the conservation of each amino acid in this region is also calculated. Exploring the restricted mutation of the V3 region could help to understand mechanism of HIV entry and to develop new strategy against HIV-1.

It is generally agreed that the recombinant gp120 (rgp120) of HIV-1 could be developed as an antibody-mediated subunit vaccine against HIV-1. Unfortunately, Connor and co-workers have provided direct experimental evidence that rgp120 (monomer) does not protect individuals from HIV-1 infection1. In addition, Moore has reviewed studies of rgp120 subunit vaccines and proposed that a new strategy is needed to develop an effective vaccine against HIV-1 (Ref. 2). An epitope vaccine might be such a strategy. The epitope vaccine against HIV-1 uses much of the principal neutralizing determinant (PND) of the envelope (Env) protein (gp160) and belongs to a special type of synthetic peptide vaccine. Several epitopes on gp160 have been characterized as PNDs, for example, GPGRAF (Env aa316-321) on the V3 loop of gp120, ELDKWA (aa669-674) on the C-domain of gp41 and RILAVERYLKD (Env aa586-596) on the N-domain of gp41. It has been demonstrated that crossneutralizing antibodies elicited by peptides of the V3 loop bind to epitope GPGRAF, which undergoes restricted mutation from GPGRAF to GPG(R/K/Q)AF (Ref. 3). ELDKWA is a relatively conserved epitope. Recent sequence analysis of primary isolates from different HIV-1 subtypes suggests that the major determinant of monoclonal antibody (mAb) 2F5 corresponds to the amino acid sequence LDKW. Naturally occurring and in vitro-selected neutralization-resistant viruses contained D to N, D to E and K to N changes in the ELDKWA motif. These amino acid changes caused abrogation of 2F5-binding to ELDKWA (Ref. 4). Recent studies have attested the breadth of reactivity of 2F5 by the antibody's significant neutralization potency against African, Asian, American and European strains from clades A, B and E. Most of the viruses investigated were neutralized by 90% (Ref. 4). The epitope RILAVERYLKD has been shown to induce protective activity5. We have designed candidate synthetic peptide vaccines and compared immunogenicity of: (1) the recombinant gp160 and gp41 subunit vaccines; (2) peptide vaccines of the C-domain on gp41 (EnvIIIB aa646-674: C-TSLIHSLIEESQNQQEKNEQELLELDKWA) and the gp120 V3 loop (EnvIIIB aa301-328: C-TRPNNNTRKSIRIQRGPGRAFYTIGKI); and (3) epitope vaccines of ELDKWA on the C-domain of gp41 [C-(ELDKWAG)4-BSA] and GPGRAFY on gp120 [C-(GPGRAFY)4-P24EC]. In mice, the rgp160 subunit vaccine induced a very weak antibody response to both epitopes (ELDKWA and GPGRAFY), whereas both synthetic peptides conjugated with carrier protein BSA or carrier peptide P24EC (GPKEPFRDYVDRFYK-C) increased the epitope-specific antibody responses (increased antibody titre by two- to fourfold) to ELDKWA and GPGRAFY. Interestingly, both the ELDKWA and the GPGRAFY tetramer epitope vaccines induced a strong epitope-specific antibody response against ELDKWA and GPGRAFY, respectively, and the levels of ELDKWA- and GPGRAFY-specific antibodies increased two- to fourfold, compared with levels induced with the synthetic peptide vaccines (Table 1). These results indicate that the epitope vaccine could provide a new strategy to develop an effective vaccine against HIV-1 infection. Crystallographic analysis indicates that the binding of gp120 to both CD4 and a chemokine receptor induces conformational changes in gp41 from the native structure to the fusion-active structure. The exposed coiled coils create a fusion intermediate, in which the C-domain and N-domain are exposed6. Recent studies have demonstrated that a fusion-competent vaccine with broad neutralization of HIV primary isolates is associated with these fusion intermediates7,8, suggesting that

nd human Iymphocytes, a highly conserved region (aa583-599; LQARILAV-ERYLKDQQL) was identified in gp41 as a binding site for human T cells and monocytes1-3, Gp41 could thus inhibit lym-phocyte proliferation4 and upregulate major histocompatibility complex (MHC) class I, class II and intercellular adhesion molecule 1 (ICAM-1) expression5-effects similar to those of human type I interferons. When we compared the sequences of gp41 and type I interferons, a similar epi-tope RILAV--YLKD was found in the im-munosuppressive domain (ISD) of gp41 and two regions in interferon α(IFN-α) (aa29-35 and 113-129), IFN-β(aa31-27 and 125-138) and IFN-ω(aa29-35 and 123-136) (Fig.1). Surprisingly, the common epitope exists within the receptor binding sites of gp41, IFN-α and IFN-β(Ref.6.) We characterized the common epitope and found increased levels of antibodies to IFN-αand IFN-βin20 HIV-infectedindividuals, compared with levels in normal healthy controls; these lev-els subsequently decreased with disease progression. Affinity-purified anti--gp41 an-tibody from HIV-infected individduals was shown to recognize human IFN-αand IFN-β. In addition, anti-IFN-αand IFN-β antibodies interacted with gp41 and its peptide(aa583-599), and inhibited gp41 binding to human lymphocytes. These results suggest that the common immunological epitope within gp41 induces incrased levels of an-tibodies that can interact with IFN-αand IFN-β. Gringeri et al. demonstrated that HIV-infected patients treated with an IFN-α vac-cine showed a significant reduction in the rate of disease progression, which was asso-ciated with an increase in IFN-αantibody titer and an increase in IFN-α-neutralizing capacity. The anti-IFN-α antibody was shown to recognize aa560-599 of HIV-1 gp41 (Ref. 7). These data suggest that the common immunological epitope within re-ceptor binding sites may be associated with protetive activity. Interestingly, a similar epitope in SIV gp32 could protect macaques from SIV infection89, indication that the common immunological epitope on gp41, IFN-αand IFN-β could be useful in the de-velopment of a vaccine strategy against HIV-infection.