Structural conservation among variants of the SARS-CoV-2 spike postfusion bundle

Yang, Kailu; Wang, Chuchu; White, K. Ian; Pfuetzner, Richard A.; Esquivies, Luis; Brunger, Axel T.

doi:10.1073/pnas.2119467119

Cited by 38 publications

(52 citation statements)

References 66 publications

(91 reference statements)

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“…For the Delta variant, the featured mutation of D950N (24410 G > A) was selected as the target site, because this mutation mapped to the trimer interface, suggesting that this mutation may contribute to the regulation of spike protein dynamics [ 39 ]. Furthermore, the Omicron variant and its subtype BA.1 contains mutation sites including N969K (24469 T > A, both Omicron and BA.1) and L981F (24503 C > T, BA.1 only) [ 40 ], and those sites were selected to distinguish them according to variants using MOPCS (Fig. 1 ).…”

Section: Resultsmentioning

confidence: 99%

A CRISPR/Cas12a-empowered surface plasmon resonance platform for rapid and specific diagnosis of the Omicron variant of SARS-CoV-2

Chen

et al. 2022

National Science Review

192

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The outbreak of the COVID-19 pandemic is partially due to the challenge of identifying asymptomatic and pre-symptomatic carriers of the virus, and thus highlights a strong motivation for diagnostics that can be rapidly deployed with high sensitivity. On the other hand, several concerned SARS-CoV-2 variants, including the Omicron, are required to be identified as soon as the samples are identified as ‘positive’. Unfortunately, a traditional PCR test does not allow their specific identification. Herein, for the first time, we have developed MOPCS (Methodologies of Photonic CRISPR Sensing), which combines an optical sensing technology-surface plasmon resonance (SPR), and the ‘gene scissors’ CRISPR technique to achieve both high sensitivity and specificity of viral variants’ measurement. MOPCS is a low-cost, CRISPR/Cas12a system-empowered SPR gene detecting platform that can analyze viral RNA, without the need for amplification, within 38 min from sample input to results output, and achieve a limit of detection of 15 fM. MOPCS achieves a highly sensitive analysis of SARS-CoV-2 and mutations appear in variants B.1.617.2 (Delta), B.1.1.529 (Omicron), and BA.1 (a subtype of Omicron). This platform was also used to analyze some recently collected patient samples from a local outbreak in China and identified by the Centers for Disease Control and Prevention. This innovative CRISPR-empowered SPR platform will further contribute to various fast, sensitive, and accurate detection of target nucleic acid sequences with single-base mutations.

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Section: Resultsmentioning

confidence: 99%

A CRISPR/Cas12a-empowered surface plasmon resonance platform for rapid and specific diagnosis of the Omicron variant of SARS-CoV-2

Chen

et al. 2022

National Science Review

192

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“…Previous high-resolution structures of the post-fusion HR1HR2 bundle of the Wuhan-Hu-1 strain of SARS-CoV-2 (GISAID accession ID: EPI_ISL_402124, here referred to as Wuh) revealed an overall six-helix bundle architecture in which the HR1 α-helices form a core with grooves into which the HR2 segments lie (PDB IDs 6lxt, 6m1v, 7rzq) (9, 15, 16). The cryo-EM structure of this HR1HR2 bundle (PDB ID 7rzq) (16) clarified several sidechain positions at high resolution and revealed that the N-terminal region of the HR2 fragment (residues 1164–1200) forms a well-defined extended conformation. Based on this observation, we further extended the N-terminal region, beginning at residue 1157, and determined the cryo-EM structure of this extended HR1HR2 complex (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The cryo-EM structure of the longHR2_45 bound to HR1 was determined following a molecular scaffolding method described previously (16). Briefly, the scaffolded HR1HR2 complex was generated by co-expressing the scaffolded HR1 and SUMO-tagged longHR2_45 in E. coli BL21(DE3) using auto-inducing LB medium (30), followed by Nickel affinity chromatography and size exclusion chromatography (SEC) with a Superose 6 Increase 10/300 GL column in 25 mM HEPES-Na, pH 7.4, 150 mM NaCl, 0.5 mM EDTA, 0.5 mM TCEP.…”

Section: Methodsmentioning

confidence: 99%

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Nanomolar inhibition of SARS-CoV-2 infection by an unmodified peptide targeting the pre-hairpin intermediate of the spike protein

Yang

Wang

Kreutzberger

et al. 2022

Preprint

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Variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) challenge currently available COVID-19 vaccines and monoclonal antibody therapies through epitope change on the receptor binding domain of the viral spike glycoprotein. Hence, there is a specific urgent need for alternative antivirals that target processes less likely to be affected by mutation, such as the membrane fusion step of viral entry into the host cell. One such antiviral class includes peptide inhibitors which block formation of the so-called HR1HR2 six-helix bundle of the SARS-CoV-2 spike (S) protein and thus interfere with viral membrane fusion. Here we performed structural studies of the HR1HR2 bundle, revealing an extended, well-folded N-terminal region of HR2 that interacts with the HR1 triple helix. Based on this structure, we designed an extended HR2 peptide that achieves single-digit nanomolar inhibition of SARS-CoV-2 in cell-based fusion, VSV-SARS-CoV-2 chimera, and authentic SARS-CoV-2 infection assays without the need for modifications such as lipidation or chemical stapling. The peptide also strongly inhibits all major SARS-CoV-2 variants to date. This extended peptide is ~100-fold more potent than all previously published short, unmodified HR2 peptides, and it has a very long inhibition lifetime after washout in virus infection assays, suggesting that it targets a pre-hairpin intermediate of the SARS-CoV-2 S protein. Together, these results suggest that regions outside the HR2 helical region may offer new opportunities for potent peptide-derived therapeutics for SARS-CoV-2 and its variants, and even more distantly related viruses, and provide further support for the pre-hairpin intermediate of the S protein.

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“…the postfusion state) relative to the bent prefusion state. [15][16][17] When this occurs away from a host cell membrane, it permanently disarms the spike trimer and also exposes more epitopes for binding of additional NAbs.…”

Section: Introductionmentioning

confidence: 99%

Identification and Mechanistic Basis of non-ACE2 Blocking Neutralizing Antibodies from COVID-19 Patients with Deep RNA Sequencing and Molecular Dynamics Simulations

Fredericks

East

Shi

et al. 2022

Preprint

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Variants of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) continue to cause disease and impair the effectiveness of treatments. The therapeutic potential of convergent neutralizing antibodies (NAbs) from fully recovered patients has been explored in several early stages of novel drugs. Here, we identified initially elicited NAbs (Ig Heavy, Ig lambda, Ig kappa) in response to COVID-19 infection in patients admitted to the intensive care unit at a single center with deep RNA sequencing (>100 million reads) of peripheral blood as a diagnostic tool for predicting the severity of the disease and as a means to pinpoint specific compensatory NAb treatments. Clinical data were prospectively collected at multiple time points during ICU admission, and amino acid sequences for the NAb CDR3 segments were identified. Patients who survived severe COVID-19 had significantly more of a Class 3 antibody (C135) to SARS-CoV-2 compared to non-survivors (16,315 reads vs 1,412 reads, p=0.02). In addition to highlighting the utility of RNA sequencing in revealing unique NAb profiles in COVID-19 patients with different outcomes, we provided a physical basis for our findings via atomistic modeling combined with molecular dynamics simulations. We established the interactions of the Class 3 NAb C135 with the SARS-CoV-2 spike protein, proposing a mechanistic basis for inhibition via multiple conformations that can effectively prevent ACE2 from binding to the spike protein, despite C135 not directly blocking the ACE2 binding motif. Overall, we demonstrate that deep RNA sequencing combined with structural modeling offers the new potential to identify and understand novel therapeutic(s) NAbs in individuals lacking certain immune responses due to their poor endogenous production. Our results suggest a possible window of opportunity for administration of such NAbs when their full sequence becomes available. A method involving rapid deep RNA sequencing of patients infected with SARS-CoV-2 or its variants at the earliest infection time could help to develop personalized treatments using the identified specific NAbs.

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Structural conservation among variants of the SARS-CoV-2 spike postfusion bundle

Cited by 38 publications

References 66 publications

A CRISPR/Cas12a-empowered surface plasmon resonance platform for rapid and specific diagnosis of the Omicron variant of SARS-CoV-2

A CRISPR/Cas12a-empowered surface plasmon resonance platform for rapid and specific diagnosis of the Omicron variant of SARS-CoV-2

Nanomolar inhibition of SARS-CoV-2 infection by an unmodified peptide targeting the pre-hairpin intermediate of the spike protein

Identification and Mechanistic Basis of non-ACE2 Blocking Neutralizing Antibodies from COVID-19 Patients with Deep RNA Sequencing and Molecular Dynamics Simulations

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