CoV-2 Spike (1000-1200)
Description
Functional Role in Viral Entry
The 1000–1200 region is critical for:
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Membrane Fusion: HR1 and CH undergo conformational changes to form a six-helix bundle with HR2, driving viral and host membrane fusion .
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Prefusion Stability: Proline substitutions (e.g., K986P, V987P) in this region stabilize the prefusion spike, enhancing vaccine immunogenicity .
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Proteolytic Activation: Cleavage at the S2' site (near residue 815) exposes the fusion peptide, enabling membrane insertion .
Glycosylation and Immune Evasion
This region hosts N1098, a glycosylation site critical for:
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Shielding Conserved Epitopes: Complex glycans at N1098 block antibody access to the fusion machinery .
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Variant Conservation: Glycosylation patterns in this region remain stable across SARS-CoV-2 variants (Alpha, Delta, Omicron), suggesting minimal immune pressure .
Research Findings and Therapeutic Implications
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Antibody Targeting: The S2 subunit (including residues 1000–1200) harbors conserved epitopes targeted by cross-reactive antibodies (e.g., CR3022), which neutralize SARS-CoV-2 by disrupting prefusion spike integrity .
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Vaccine Design: Stabilizing mutations (e.g., D614G) in the S2 subunit improve spike trimer stability, enhancing vaccine efficacy .
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Neurotropism: Spike protein fragments, including S2 residues, persist in the skull marrow and brain parenchyma post-infection, potentially contributing to long-term neurological symptoms .
Comparative Analysis of Spike Domains
Key Mutations and Variant Impact
While most mutations in circulating variants (e.g., Delta, Omicron) occur in S1, the AY.1 (Delta plus) variant retains critical S2 features (e.g., intact HR1 and glycosylation sites), preserving fusion functionality and immune evasion .
Product Specs
In December 2019, a new coronavirus, known as 2019 novel coronavirus (2019-nCoV), emerged in Wuhan, China. This virus, responsible for causing viral pneumonia in humans, was first identified in a seafood market.
Genetic analysis revealed that 2019-nCoV shares a significant similarity (87% identity) with the bat-derived SARS-CoV-2, discovered in Zhoushan, eastern China, in 2018. Despite some differences, the receptor-binding domain (RBD) structure of 2019-nCoV closely resembles that of 2018 SARS-CoV, suggesting its potential to bind to the human ACE2 receptor (angiotensin-converting enzyme 2).
While bats are considered the likely natural reservoir of 2019-nCoV, it is suspected that an intermediary animal host, possibly one traded at the Wuhan market, played a role in its transmission to humans. Research indicates that 2019-nCoV might have arisen from a recombination event involving the spike glycoprotein of a bat coronavirus and another, yet unidentified, coronavirus.
This recombinant protein, produced in E. coli, encompasses the immunodominant regions of the Coronavirus 2019 Spike protein (amino acids 1000-1200). It is equipped with a C-terminal 6xHis tag for purification and detection purposes.
The CoV 2019 Spike Protein is provided as a 1 mg/ml solution in 1x PBS (phosphate-buffered saline).
To maintain product integrity, CoV 2019 Spike Protein is shipped using ice packs. Upon receipt, it should be stored at -20°C.
The purity of the CoV 2019 Spike Protein exceeds 90%, as assessed by SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis).
Q&A
Basic Research Questions
What experimental strategies optimize structural analysis of the Spike protein’s 1000-1200 region?
Combine cryo-EM for global conformational states (>3.5 Å resolution) with molecular dynamics (MD) simulations (all-atom structure-based models) to capture steric effects and glycan interactions . For example:
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Use biased MD simulations initiated from prefusion conformations to study S2’ subunit rearrangements
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Employ surface plasmon resonance (SPR) and bio-layer interferometry (BLI) to quantify binding kinetics (Table 1)
How does the 1000-1200 region influence ACE2 binding stability?
This region facilitates a secondary conformational state post-initial RBD-ACE2 contact:
| Parameter | Primary Binding (Langmuir) | Secondary State Transition | Total Affinity |
|---|---|---|---|
| Association Rate | 1.8×10⁵ M⁻¹s⁻¹ | 8.3×10⁻⁴ s⁻¹ | - |
| Dissociation Rate | 6.7×10⁻³ s⁻¹ | 8.5×10⁻⁵ s⁻¹ | 0.20 nM |
The secondary state increases complex half-life by 80× through structural rigidification . Validate using trimeric spike constructs to observe cooperative stabilization effects .
Advanced Research Challenges
What mutagenesis approaches best test fusion mechanism hypotheses in this region?
Use deep mutational scanning combined with pseudovirus assays:
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Introduce H519N substitution to study replication deficits in lung epithelium
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Pair with hydrogen-deuterium exchange MS to map allosteric effects
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Benchmark against ancestral strain using evolutionary analyses (OmegaPlus pipeline)
Why do glycosylation patterns in this region show context-dependent effects on fusion?
MD simulations reveal glycan steric effects create a kinetic trap:
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Glycan density >60% extends intermediate state lifetime by 2.3×
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N-linked glycans at N1074 act as conformational "ratchets" during S1/S2 separation
Validate using glycoengineered pseudoviruses with defined glycoforms .
Data Conflict Resolution Framework
Product Science Overview
The spike protein is a large glycoprotein that protrudes from the surface of the SARS-CoV-2 virus. It is composed of two subunits, S1 and S2, which are responsible for different functions:
- S1 Subunit: Contains the receptor-binding domain (RBD) that binds to the angiotensin-converting enzyme 2 (ACE2) receptor on the surface of host cells.
- S2 Subunit: Facilitates the fusion of the viral membrane with the host cell membrane, allowing the viral genome to enter the host cell.
The spike protein is synthesized as a precursor that undergoes proteolytic cleavage to become functional. The segment from amino acids 1000 to 1200 is part of the S2 subunit, which is critical for the membrane fusion process.
Recombinant proteins are produced through recombinant DNA technology, which involves inserting the gene encoding the protein into an expression system, such as bacteria, yeast, or mammalian cells. The expression system then produces the protein, which can be purified for various applications.
The recombinant spike protein segment (1000-1200 a.a.) is used in research and vaccine development for several reasons:
- Immunogenicity: This segment contains epitopes that can elicit an immune response, making it a valuable target for vaccine development.
- Structural Studies: Understanding the structure of this segment helps in elucidating the mechanisms of viral entry and fusion.
- Diagnostic Tools: Recombinant spike proteins are used in serological assays to detect antibodies against SARS-CoV-2 in patient samples.
The spike protein, including the 1000-1200 a.a. segment, is a key antigen in many COVID-19 vaccines. For example, the NVX-CoV2373 vaccine developed by Novavax uses a recombinant spike protein nanoparticle to induce an immune response . This vaccine has shown promising results in clinical trials, demonstrating strong immunogenicity and protection against COVID-19.
