Recombinant HIV type 1 Glycoprotein 41 (a.a. 541-682) [His]

Many broadly neutralizing antibodies (bnAbs) against HIV-1 recognize and/or penetrate the glycan shield on native, virion-associated envelope glycoprotein (Env) spikes. The same bnAbs also bind to recombinant, soluble trimeric immunogens based on the SOSIP design. While SOSIP trimers are close structural and antigenic mimics of virion Env, the extent to which their glycan structures resemble ones on infectious viruses is undefined. Here, we compare the overall glycosylation of gp120 and gp41 subunits from BG505 (clade A) virions produced in a lymphoid cell line with those from recombinant BG505 SOSIP trimers, including CHO-derived clinical grade material. We also performed detailed site-specific analyses of gp120. Glycans relevant to key bnAb epitopes are generally similar on the recombinant SOSIP and virion-derived Env proteins, although the latter do contain hotspots of elevated glycan processing. Knowledge of native versus recombinant Env glycosylation will guide vaccine design and manufacturing programs.

To characterize HIV-1 gp41 as an antigen for developing HIV-1 incidence assay and to investigate the impact of HIV-1 genetic diversity on the assay performance, a number of truncated peptides were synthesized to identify the immunodominant epitopes (IDEs) of HIV-1 gp41 protein. Subsequently, the mixed peptides (MP3) or the recombinant protein (MP4) containing HIV-1 gp41 IDEs of the major HIV-1 genotype CRF01_AE, CRF07_BC/CRF08_BC and subtype B in China were used to verify the sensitivity and specificity of HIV-1 recency testing. We identified the QKFLG and GKIIC motifs located in the loop region of HIV-1 gp41 as the two major IDEs. The surrounding amino acids EAQQHLLQLT and WNSSWSN could block the binding of gp41 peptide and anti-HIV antibody with low avidity, making the gp41 peptide p57 suitable for distinguishing recent and long-term HIV-1 infections. Furthermore, MP3 or MP4-based immunoassay could significantly improve the assay sensitivity and showed 93.33% (140/150) vs. 94.59% (35/37) and 94.08% (143/152) vs. 94.59% (35/37) concordance with commercially available LAg-Avidity EIA test among the cross-sectional and longitudinal samples, respectively. The estimated mean duration of recent infection (MDRI) was 130 days (95% CI: 83-167) and 166 days (95% CI: 123-202) for MP3 and MP4 assays, respectively. Our preliminary results indicate that the HIV-1 gp41 peptide-based immunoassay specifically targeting the major HIV-1 genotype CRF01_AE, CRF07_BC/CRF08_BC and subtype B could serve as a simple incidence assay for differentiating recent and long-term HIV-1 infections in China.