Abstract
To provide protective immunity against circulating primary HIV-1 strains, a vaccine most likely has to induce broadly neutralizing antibodies to the HIV-1 envelope glycoprotein (Env) spike. Recombinant Env trimers such as the prototype BG505 SOSIP.664 that closely mimic the native Env spike can induce autologous neutralizing antibodies (NAbs) against relatively resistant (tier 2) primary viruses. Ideally, Env immunogens should present broadly neutralizing antibody epitopes but limit the presentation of immunodominant non-NAb epitopes that might induce off-target and potentially interfering responses. The V3 loop in gp120 is such a non-NAb epitope that can effectively elicit non-NAbs when animals are immunized with SOSIP.664 trimers. V3 immunogenicity can be diminished, but not abolished, by reducing the conformational flexibility of trimers via targeted sequence changes, including an A316W substitution in V3, that create the SOSIP.v4.1 and SOSIP.v5.2 variants. Here, we further modified these trimer designs by introducing leucine residues at V3 positions 306 and 308 to create hydrophobic interactions with the tryptophan residue at position 316 and with other topologically proximal sites in the V1V2 domain. Together, these modifications further stabilized the resulting SOSIP.v5.2 S306L/R308L trimers in the prefusion state in which V3 is sequestered. When we tested these trimers as immunogens in rabbits, the induction of V3 non-NAbs was significantly reduced compared with the SOSIP.v5.2 trimers and even more so compared with the SOSIP.664 prototype, without affecting the autologous NAb response. Hence, these additional trimer sequence modifications may be beneficial for immunization strategies that seek to minimize off-target non-NAb responses.
Highlights
Introduction of hydrophobic residues inV3 reduces envelope glycoprotein (Env) functionTo determine the effect of the S306L and R308L substitutions on Env function, we introduced them into the infectious BG505.T332N Env-pseudotyped virus and measured infectivity on TZM-bl cells (Fig. 2)
To strengthen the hydrophobic interactions that maintain the burial of the V3 region in the pocket formed by the adjacent V1V2 elements, we introduced leucine residues at V3 positions 306 and 308 (Fig. 1, A–D)
The various designs and genotypes of native-like SOSIP trimers serve as a platform for the eventual induction of broadly neutralizing antibodies (bNAbs), which is a challenging task that will require manipulating the immunogen design so as to reproduce consistently how bNAbs are sometimes elicited during HIV-1 infection [49, 50]
Summary
We previously found that the introduction of an A316W mutation into V3 reduced the exposure of this Env domain on the trimer surface and thereby reduced the induction of V3-directed non-NAbs by ϳ10-fold when the resulting SOSIP.v4 trimers were tested as immunogens in mice and rabbits, but to a lesser extent in macaques [25, 41]. The V3 crown predominantly participates in intraprotomer hydrophobic interactions with the V1V2 loop and with the N-terminal region of the V1V2 stem of the same protomer [44] Disrupting these cross-domain hydrophobic interactions changes the packing of the V3 region, destabilizes its location in the pocket, and triggers a transition toward alternative, energetically favorable conformations in which V3 is surface-exposed [44]. To strengthen the hydrophobic interactions that maintain the burial of the V3 region in the pocket formed by the adjacent V1V2 elements, we introduced leucine residues at V3 positions 306 and 308 (Fig. 1, A–D). We used PyMOL to identify the residues in contact with (distance of Ͻ4 or Ͻ5 Å) amino acids Ser306, Arg308, and Ala316 in the structure of the wild-type BG505 SOSIP.664 trimer (Protein Data Bank code 5CEZ) [93] and employed an in silico model to assess the contact residues for the hydrophobic substitutions at these positions, i.e., Leu306, Leu308, and Trp316 [94]. Residues 306, 308, and 316 are highlighted in bold type
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