Structure-Based Stabilization of SOSIP Env Enhances Recombinant Ectodomain Durability and Yield

Wrapp, Daniel; Mu, Zekun; Thakur, Bhishem; Janowska, Katarzyna; Ajayi, Oluwatobi; Barr, Maggie; Parks, Robert; Mansouri, Katayoun; Edwards, Robert J.; Hahn, Beatrice H.; Acharya, Priyamvada; Saunders, Kevin O.; Haynes, Barton F.

doi:10.1128/jvi.01673-22

Cited by 13 publications

(9 citation statements)

References 52 publications

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“…Examination of a representative set of State 3 structures suggested that replacing the N386-glycan with an arginine could aid in attracting this state through interaction with CH235.12 F v aspartate and glutamate residues (Supplemental Figure 21B). Consistent with Encounter State 3 structural observations, an N386A mutation, referred to here as CH235 Encounter design 0 (CE0), enhanced the CH235.12 Fab association rate by ~2 fold and a stabilized form of an N386R mutation, referred to here as CE1, which contains the F14 and 2P stabilization mutations (44,45), enhanced the CH235.12 Fab by ~6 fold (Figure 4E, Supplemental Figures 23-25 and 26B, Supplemental Table 2). The non-stabilized CE1 design showed only ~2-fold enhanced association rate, suggesting stabilization affected the rate (Supplemental Table 2).…”

Section: Association Pathway-designed Immunogens For Bnab Immunogens ...supporting

confidence: 68%

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Engineering immunogens that select for specific mutations in HIV broadly neutralizing antibodies

Henderson,

Anasti,

Manne

et al. 2023

Preprint

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Vaccine development targeting rapidly evolving pathogens such as HIV-1 requires induction of broadly neutralizing antibodies (bnAbs) with conserved paratopes and mutations, and, in some cases, the same Ig-heavy chains. The current trial-and-error search for immunogen modifications that improve selection for specific bnAb mutations is imprecise. To precisely engineer bnAb boosting immunogens, we used molecular dynamics simulations to examine encounter states that form when antibodies collide with the HIV-1 Envelope (Env). By mapping how bnAbs use encounter states to find their bound states, we identified Env mutations that were predicted to select for specific antibody mutations in two HIV-1 bnAb B cell lineages. The Env mutations encoded antibody affinity gains and selected for desired antibody mutations in vivo. These results demonstrate proof-of-concept that Env immunogens can be designed to directly select for specific antibody mutations at residue-level precision by vaccination, thus demonstrating the feasibility of sequential bnAb-inducing HIV-1 vaccine design.

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Section: Association Pathway-designed Immunogens For Bnab Immunogens ...supporting

confidence: 68%

“…The DS stabilized ( 58 ) CH848.10.17.d949 DT and DE1-3 envelope SOSIPs and the 4.1 ( 59 ), 2P ( 45 ), and F14 stabilized ( 44 ) CH505.M5.G458Y, CE0-3 SOSIPs were expressed in Freestyle TM 293-F cells (ThermoFisher Cat No. R79007).…”

Section: Methodsmentioning

confidence: 99%

Engineering immunogens that select for specific mutations in HIV broadly neutralizing antibodies

Henderson,

Anasti,

Manne

et al. 2023

Preprint

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“…From this analysis, we identified a V127C-D167C disulfide that stapled V1/V2 contact regions between gp120 protomers in a CH505 SOSIP trimer design parent ( 58 ). This design was further stabilized using the previously reported F14 ( 39 ) and SOSIP 2P ( 59 ) mutations to improve expression and folding in addition to a CD4bs antibody binding enhancing N197D glycan deletion mutation ( 60 ). A closed Env trimer conformation was identified by negative-stain electron microscopy (NSEM).…”

Section: Resultsmentioning

confidence: 99%

Microsecond dynamics control the HIV-1 Envelope conformation

Bennett,

Edwards,

Kosheleva

et al. 2024

Sci. Adv.

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The HIV-1 Envelope (Env) glycoprotein facilitates host cell fusion through a complex series of receptor-induced structural changes. Although remarkable progress has been made in understanding the structures of various Env conformations, microsecond timescale dynamics have not been studied experimentally. Here, we used time-resolved, temperature-jump small-angle x-ray scattering to monitor structural rearrangements in an HIV-1 Env SOSIP ectodomain construct with microsecond precision. In two distinct Env variants, we detected a transition that correlated with known Env structure rearrangements with a time constant in the hundreds of microseconds range. A previously unknown structural transition was also observed, which occurred with a time constant below 10 μs, and involved an order-to-disorder transition in the trimer apex. Using this information, we engineered an Env SOSIP construct that locks the trimer in the prefusion closed state by connecting adjacent protomers via disulfides. Our findings show that the microsecond timescale structural dynamics play an essential role in controlling the Env conformation with impacts on vaccine design.

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“…Since then, a plethora of stabilized Envs from different HIV-1 clades have been reported. They are all based on the SOSIP.664 design and include additional disulfides, cavity filling mutations, and other proline mutations, all leading to enhanced thermostability of the Env trimers [ 94 , 95 , 96 , 97 , 98 , 99 , 100 , 101 , 102 ].…”

Section: Viral Glycoprotein Stabilization Strategiesmentioning

confidence: 99%

SARS-CoV-2 S Glycoprotein Stabilization Strategies

Pedenko

Sulbarán

Guilligay

et al. 2023

Viruses

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The SARS-CoV-2 pandemic has again shown that structural biology plays an important role in understanding biological mechanisms and exploiting structural data for therapeutic interventions. Notably, previous work on SARS-related glycoproteins has paved the way for the rapid structural determination of the SARS-CoV-2 S glycoprotein, which is the main target for neutralizing antibodies. Therefore, all vaccine approaches aimed to employ S as an immunogen to induce neutralizing antibodies. Like all enveloped virus glycoproteins, SARS-CoV-2 S native prefusion trimers are in a metastable conformation, which primes the glycoprotein for the entry process via membrane fusion. S-mediated entry is associated with major conformational changes in S, which can expose many off-target epitopes that deviate vaccination approaches from the major aim of inducing neutralizing antibodies, which mainly target the native prefusion trimer conformation. Here, we review the viral glycoprotein stabilization methods developed prior to SARS-CoV-2, and applied to SARS-CoV-2 S, in order to stabilize S in the prefusion conformation. The importance of structure-based approaches is highlighted by the benefits of employing stabilized S trimers versus non-stabilized S in vaccines with respect to their protective efficacy.

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Structure-Based Stabilization of SOSIP Env Enhances Recombinant Ectodomain Durability and Yield

Cited by 13 publications

References 52 publications

Engineering immunogens that select for specific mutations in HIV broadly neutralizing antibodies

Engineering immunogens that select for specific mutations in HIV broadly neutralizing antibodies

Microsecond dynamics control the HIV-1 Envelope conformation

SARS-CoV-2 S Glycoprotein Stabilization Strategies

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