CoV-2 S1 (319-537)
Description
Definition and Context
CoV-2 S1 (319-537) refers to a recombinant protein fragment derived from the receptor-binding domain (RBD) of the SARS-CoV-2 spike (S) glycoprotein. This region spans amino acids 319–537 within the S1 subunit and is critical for viral entry into host cells via interaction with the human angiotensin-converting enzyme 2 (ACE2) receptor . It is widely studied for vaccine development, therapeutic antibody design, and serological diagnostics due to its immunodominant epitopes .
Domain Architecture
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S1/S2 Subunits: The SARS-CoV-2 spike protein is divided into S1 (receptor-binding) and S2 (membrane fusion) subunits. The S1 subunit contains the RBD (residues 319–541), which adopts a β-sheet core stabilized by disulfide bonds .
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Key Residues:
Conformational Dynamics
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The RBD exists in "open" (ACE2-accessible) and "closed" states, with structural flexibility enabling immune evasion .
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Trimerization of RBD (tri-RBD) mimics native spike arrangements, improving immunogenicity compared to monomeric forms .
Antigenicity and Neutralization
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CoV-2 S1 (319-537) contains neutralizing epitopes targeted by >90% of antibodies in convalescent sera .
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Key Findings:
Cross-Reactivity
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Antibodies against this region show partial cross-neutralization of variants (e.g., Beta, Delta, Omicron) but require updates for emerging strains .
Vaccine Development
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Prototype-Beta Chimeric RBD-Dimer: Elicited broad protection in macaques, with neutralizing titers of 1:512 against Omicron .
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HEK293-6E Cell Expression: Optimized RBD (319–541) yields 20–25% dimers, necessitating C538 mutations (e.g., C538A) to stabilize monomers .
Therapeutic Targeting
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Thiol-Based Drugs: Cysteamine and WR-1065 disrupt the Cys480-Cys488 disulfide bond, reducing ACE2 binding by >80% .
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Furin Cleavage Site (FCS): While absent in CoV-2 S1 (319-537), FCS engineering in related constructs enhances cell-cell fusion and infectivity .
Challenges in Production and Stability
Comparative Analysis with SARS-CoV-1
Future Directions
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Pan-Coronavirus Vaccines: Hybrid RBD designs incorporating mutations from multiple variants (e.g., Delta-Omicron chimera) .
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Adjuvant Optimization: Screening novel adjuvants to enhance RBD-specific T-cell responses .
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Structural Biology: Cryo-EM studies to map conformational changes during ACE2 binding .
Product Specs
The novel coronavirus (2019-nCoV) responsible for the 2019 outbreak of viral pneumonia was identified in December 2019 in Wuhan, China. This virus, initially found in a seafood market, shares a high genetic similarity with bat coronaviruses, suggesting a possible animal origin.
Genetic analysis reveals that 2019-nCoV shares 87% identity with bat SARS-related coronaviruses, specifically the SARS-CoV-2 strain. The virus's receptor-binding domain (RBD) structure closely resembles that of SARS-CoV, enabling it to potentially bind to the human ACE2 receptor, a key protein involved in regulating blood pressure.
While bats are suspected to be the natural reservoir of 2019-nCoV, an intermediate animal host, potentially from the seafood market, is thought to have facilitated transmission to humans. Notably, research suggests that 2019-nCoV's spike glycoprotein, crucial for host cell entry, might be a product of recombination between a bat coronavirus and an as-yet-unidentified coronavirus.
This recombinant protein, derived from HEK293 cells, corresponds to the Receptor Binding Domain (RBD) of the SARS-CoV-2 Spike Glycoprotein S1 (Wuhan-Hu-1 strain, amino acids 319-537). This protein, with a molecular weight of 26.5kDa, includes a C-terminal His tag for purification and detection.
The product is lyophilized (freeze-dried) from a solution containing PBS at pH 7.4, 10% trehalose, and has been sterile filtered through a 0.2µm filter.
For optimal reconstitution, dissolve the lyophilized CoV-2 S1 protein in sterile, ultrapure water (18 MΩ·cm) to a concentration of 0.5 mg/ml. A minimum concentration of 0.1 mg/ml is acceptable. The reconstituted protein can be further diluted in other aqueous solutions as needed.
While the lyophilized Cov-2 RBD protein remains stable at room temperature for up to three weeks, it is recommended to store it desiccated at a temperature below -18°C. After reconstitution, the CoV2 RBD protein can be stored at 4°C for 2-7 days. For long-term storage, freeze the reconstituted protein below -18°C. It is advisable to add a carrier protein (0.1% HSA or BSA) to enhance stability during storage. Avoid repeated freeze-thaw cycles.
The purity of this protein is determined to be greater than 90% using SDS-PAGE analysis.
HEK293 Cells.
Purified by Metal-Afinity chromatographic technique.
Q&A
What is the functional significance of the S1 (319-537) region in SARS-CoV-2 pathogenesis?
The S1 (319-537) region forms the core of the RBD, mediating viral attachment to angiotensin-converting enzyme 2 (ACE2) receptors on host cells. Structural studies using cryo-EM and molecular dynamics simulations reveal that residues like N501, Y453, and K417 directly participate in ACE2 binding through hydrophobic and electrostatic interactions . Methodologically, surface plasmon resonance (SPR) and ELISA are standard for quantifying RBD-ACE2 binding kinetics (e.g., dissociation constants ranging from 1.2 nM to 15 nM across variants) . Researchers should note that pseudovirus neutralization assays are critical for validating RBD-antibody interactions in vitro .
Which experimental techniques are optimal for characterizing S1 (319-537) structural dynamics?
Atomic force microscopy (AFM) has proven effective for measuring unbinding forces between RBD and host receptors. For example, studies report unbinding forces of 75–120 pN for RBD-ACE2 interactions and 90–130 pN for RBD-neuropilin-1 (NRP1) interactions, suggesting NRP1’s role as a co-receptor . Comparative circular dichroism spectroscopy further reveals that the RBD maintains β-sheet dominance (55–60% of secondary structure) even under mutagenesis . For computational validation, molecular dynamics simulations with AMBER or CHARMM force fields are recommended, particularly for analyzing variant-induced conformational changes .
Table 1: Biophysical Techniques for RBD Analysis
Why do clinical studies report conflicting data on RBD persistence and post-acute sequelae?
A 2024 longitudinal cohort study detected serum RBD in 14% of post-COVID ME/CFS patients but found no correlation with symptom severity (ρ = -0.12, p = 0.24) . This contrasts with tissue-based studies showing RBD persistence in 30% of fatal COVID-19 cases. Methodological factors explain discrepancies:
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ELISA sensitivity thresholds: Commercial assays detect RBD at ≥10 pg/mL, missing immune-complexed spike .
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Compartmentalization: Serum vs. tissue RBD pools may reflect distinct clearance mechanisms .
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Temporal dynamics: RBD clearance correlates weakly with time post-infection (r = 0.18, p = 0.07) .
Researchers should combine ultrasensitive immunoassays (e.g., Simoa) with lymphocyte profiling to resolve these contradictions.
Table 2: Cohort Studies of RBD Persistence
Can computational models reliably predict RBD mutation impacts on transmissibility?
The empirical scoring function (ESF) model, trained on 281,000+ binding energy measurements, predicts variant-specific ACE2 affinities with ±2 kJ/mol accuracy . Key findings:
Methodological Recommendations
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For binding kinetics: Use AFM at 0.5–1.0 nN/s loading rates to mimic physiological shear forces .
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For variant analysis: Pair ESF models with alanine-scanning mutagenesis to identify epistatic residues .
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For clinical studies: Employ immunoglobulin depletion (e.g., protein A/G columns) to detect immune-complexed RBD .
Product Science Overview
The Coronavirus 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has had a profound impact on global health. A critical component of the virus is the spike (S) glycoprotein, which plays a pivotal role in the virus’s ability to infect host cells. The spike glycoprotein is composed of two subunits, S1 and S2, with the S1 subunit containing the receptor-binding domain (RBD) that is essential for the virus to attach to and enter host cells .
The spike glycoprotein is a large type I transmembrane protein that protrudes from the surface of the virus. The S1 subunit, specifically the receptor-binding domain (RBD), spans amino acids 319 to 537. This domain is responsible for binding to the angiotensin-converting enzyme 2 (ACE2) receptor on the surface of host cells . The interaction between the RBD and ACE2 is the initial step in the viral entry process, facilitating the fusion of the viral and cellular membranes .
Recombinant production of the SARS-CoV-2 spike glycoprotein S1 RBD involves expressing the protein in a suitable host system, such as mammalian cells, insect cells, or yeast. This allows for the production of large quantities of the protein for research and therapeutic purposes. The recombinant S1 RBD can be used in various applications, including vaccine development, diagnostic assays, and therapeutic interventions .
The S1 RBD is a major target for neutralizing antibodies, making it a key focus for vaccine development. Vaccines that elicit an immune response against the RBD can potentially block the virus from binding to the ACE2 receptor, thereby preventing infection. Several COVID-19 vaccines, including mRNA-based vaccines, have been designed to induce an immune response against the spike glycoprotein, particularly the RBD .
In addition to its role in vaccine development, the recombinant S1 RBD is used in therapeutic and diagnostic applications. For instance, it can be employed in serological assays to detect antibodies against SARS-CoV-2 in patient samples. Furthermore, the RBD can be used to screen for potential antiviral compounds that inhibit the interaction between the virus and the ACE2 receptor .
