scholarly journals Characterization of the SARS-CoV-2 Spike in an Early Prefusion Conformation

2020 ◽  
Author(s):  
Tingting Li ◽  
Qingbing Zheng ◽  
Hai Yu ◽  
Dinghui Wu ◽  
Wenhui Xue ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Severe Acute Respiratory Syndrome ◽  
Receptor Binding ◽  
Insect Cells ◽  
Vaccine Development ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Viral Membrane ◽  
Cryo Electron Microscopy

AbstractPandemic coronavirus disease 2019 (COVID-19) is caused by the emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), for which there are no efficacious vaccines or therapeutics that are urgently needed. We expressed three versions of spike (S) proteins—receptor binding domain (RBD), S1 subunit and S ectodomain—in insect cells. RBD appears monomer in solutions, whereas S1 and S associate into homotrimer with substantial glycosylation. The three proteins confer excellent antigenicity with six convalescent COVID-19 patient sera. Cryo-electron microscopy (cryo-EM) analyses indicate that the SARS-CoV-2 S trimer dominate in a unique conformation distinguished from the classic prefusion conformation of coronaviruses by the upper S1 region at lower position ~15 Å proximal to viral membrane. Such conformation is proposed as an early prefusion state for the SARS-CoV-2 spike that may broaden the knowledge of coronavirus and facilitate vaccine development.

scholarly journals Potent Neutralization of SARS-CoV-2 by Hetero-Bivalent Alpaca Nanobodies Targeting the Spike Receptor-Binding Domain

2021 ◽  
Vol 95 (10) ◽  
Author(s):  
Huan Ma ◽  
Weihong Zeng ◽  
Xiangzhi Meng ◽  
Xiaoxue Huang ◽  
Yunru Yang ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Vaccine Development ◽  
Human Life ◽  
Pulmonary Delivery ◽  
Binding Domain ◽  
Binding Affinities ◽  
Receptor Binding Domain ◽  
Angiotensin Converting Enzyme 2 ◽  
Economic Output

ABSTRACT Cellular entry of SARS-CoV-2 requires the binding between the receptor-binding domain (RBD) of the viral Spike protein and the cellular angiotensin-converting enzyme 2 (ACE2). As such, RBD has become the major target for vaccine development, whereas RBD-specific antibodies are pursued as therapeutics. Here, we report the development and characterization of SARS-CoV-2 RBD-specific VHH/nanobody (Nb) from immunized alpacas. Seven RBD-specific Nbs with high stability were identified using phage display. They bind to SARS-CoV-2 RBD with affinity KD ranging from 2.6 to 113 nM, and six of them can block RBD-ACE2 interaction. The fusion of the Nbs with IgG1 Fc resulted in homodimers with greatly improved RBD-binding affinities (KD ranging from 72.7 pM to 4.5 nM) and nanomolar RBD-ACE2 blocking abilities. Furthermore, the fusion of two Nbs with nonoverlapping epitopes resulted in hetero-bivalent Nbs, namely, aRBD-2-5 and aRBD-2-7, with significantly higher RBD binding affinities (KD of 59.2 pM and 0.25 nM) and greatly enhanced SARS-CoV-2 neutralizing potency. The 50% neutralization doses of aRBD-2-5 and aRBD-2-7 were 1.22 ng/ml (∼0.043 nM) and 3.18 ng/ml (∼0.111 nM), respectively. These high-affinity SARS-CoV-2 blocking Nbs could be further developed into therapeutics, as well as diagnostic reagents for COVID-19. IMPORTANCE To date, SARS-CoV-2 has caused tremendous loss of human life and economic output worldwide. Although a few COVID-19 vaccines have been approved in several countries, the development of effective therapeutics, including SARS-CoV-2 targeting antibodies, remains critical. Due to their small size (13 to 15 kDa), high solubility, and stability, Nbs are particularly well suited for pulmonary delivery and more amenable to engineer into multivalent formats than the conventional antibody. Here, we report a series of new anti-SARS-CoV-2 Nbs isolated from immunized alpaca and two engineered hetero-bivalent Nbs. These potent neutralizing Nbs showed promise as potential therapeutics against COVID-19.

scholarly journals Implications for SARS-CoV-2 Vaccine Design: Fusion of Spike Glycoprotein Transmembrane Domain to Receptor-Binding Domain Induces Trimerization

Membranes ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 215 ◽  
Author(s):  
Taha Azad ◽  
Ragunath Singaravelu ◽  
Mathieu J.F. Crupi ◽  
Taylor Jamieson ◽  
Jaahnavi Dave ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Transmembrane Domain ◽  
Vaccine Development ◽  
Binding Domain ◽  
Effective Vaccine ◽  
Therapeutic Drug ◽  
Receptor Binding Domain ◽  
Design Strategies ◽  
Protein Receptor

The ongoing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic presents an urgent need for an effective vaccine. Molecular characterization of SARS-CoV-2 is critical to the development of effective vaccine and therapeutic strategies. In the present study, we show that the fusion of the SARS-CoV-2 spike protein receptor-binding domain to its transmembrane domain is sufficient to mediate trimerization. Our findings may have implications for vaccine development and therapeutic drug design strategies targeting spike trimerization. As global efforts for developing SARS-CoV-2 vaccines are rapidly underway, we believe this observation is an important consideration for identifying crucial epitopes of SARS-CoV-2.

scholarly journals Structure of Severe Acute Respiratory Syndrome Coronavirus Receptor-binding Domain Complexed with Neutralizing Antibody

2006 ◽  
Vol 281 (23) ◽  
pp. 15829-15836 ◽  
Author(s):  
Ponraj Prabakaran ◽  
Jianhua Gan ◽  
Yang Feng ◽  
Zhongyu Zhu ◽  
Vidita Choudhry ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Receptor Binding ◽  
Neutralizing Antibody ◽  
Binding Domain ◽  
Receptor Binding Domain

scholarly journals Cross-reactive neutralization of SARS-CoV-2 by serum antibodies from recovered SARS patients and immunized animals

2020 ◽  
Vol 6 (45) ◽  
pp. eabc9999 ◽  
Author(s):  
Yuanmei Zhu ◽  
Danwei Yu ◽  
Yang Han ◽  
Hongxia Yan ◽  
Huihui Chong ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Receptor Binding ◽  
Binding Domain ◽  
Full Length ◽  
Receptor Binding Domain ◽  
Serum Antibodies ◽  
Serum Samples ◽  
S Protein ◽  
Novel Coronavirus

The current coronavirus disease 2019 (COVID-19) pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel coronavirus genetically close to SARS-CoV. To investigate the effects of previous SARS-CoV infection on the ability to recognize and neutralize SARS-CoV-2, we analyzed 20 convalescent serum samples collected from individuals infected with SARS-CoV during the 2003 SARS outbreak. All patient sera reacted strongly with the S1 subunit and receptor binding domain (RBD) of SARS-CoV; cross-reacted with the S ectodomain, S1, RBD, and S2 proteins of SARS-CoV-2; and neutralized both SARS-CoV and SARS-CoV-2 S protein–driven infections. Analysis of antisera from mice and rabbits immunized with a full-length S and RBD immunogens of SARS-CoV verified cross-reactive neutralization against SARS-CoV-2. A SARS-CoV–derived RBD from palm civets elicited more potent cross-neutralizing responses in immunized animals than the RBD from a human SARS-CoV strain, informing strategies for development of universal vaccines against emerging coronaviruses.

Receptor-binding domain of human α2-macroglobulin Expression, folding and biochemical characterization of a high-affinity recombinant derivative

FEBS Letters ◽  
1994 ◽  
Vol 344 (2-3) ◽  
pp. 242-246 ◽  
Author(s):  
Thor Las Holtet ◽  
Kåre Lehmann Nielsen ◽  
Michael Etzerodt ◽  
Søren Kragh Moestrup ◽  
Jørgen Gliemann ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Biochemical Characterization ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
High Affinity ◽  
Α2 Macroglobulin

scholarly journals An antibody-escape calculator for mutations to the SARS-CoV-2 receptor-binding domain

2021 ◽  
Author(s):  
Allison J Greaney ◽  
Tyler N Starr ◽  
Jesse D Bloom
Keyword(s):  
Receptor Binding ◽  
Polyclonal Antibody ◽  
Polyclonal Antibodies ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Experimental Characterization ◽  
Antibody Recognition ◽  
Antigenic Properties ◽  
Interactive Version

A key goal of SARS-CoV-2 surveillance is to rapidly identify viral variants with mutations that reduce neutralization by polyclonal antibodies elicited by vaccination or infection. Unfortunately, direct experimental characterization of new viral variants lags their sequence-based identification. Here we help address this challenge by aggregating deep mutational scanning data into an "escape calculator" that estimates the antigenic effects of arbitrary combinations of mutations to the virus's spike receptor-binding domain (RBD). The calculator can be used to intuitively visualize how mutations impact polyclonal antibody recognition, and score the expected antigenic effect of combinations of mutations. These scores correlate with neutralization assays performed on SARS-CoV-2 variants, and emphasize the ominous antigenic properties of the recently described Omicron variant. An interactive version of the calculator is at https://jbloomlab.github.io/SARS2_RBD_Ab_escape_maps/escape-calc/, and we provide a Python module for batch processing.

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