scholarly journals The SARS-CoV-2 Spike Variant D614G Favors an Open Conformational State

2020 ◽  
Author(s):  
Rachael A. Mansbach ◽  
Srirupa Chakraborty ◽  
Kien Nguyen ◽  
David C. Montefiori ◽  
Bette Korber ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Molecular Level ◽  
Receptor Binding Domain ◽  
Target Region ◽  
Infected People ◽  
Viral Loads ◽  
Rapid Transition ◽  
Open States ◽  
Insight Into

SummaryThe COVID-19 pandemic underwent a rapid transition with the emergence of a SARS-CoV-2 variant that carried the amino acid substitution D614G in the Spike protein that became globally prevalent. The G-form is both more infectious in vitro and associated with increased viral loads in infected people. To gain insight into the mechanism underlying these distinctive characteristics, we employed multiple replicas of microsecond all-atom simulations to probe the molecular-level impact of this substitution on Spike’s closed and open states. The open state enables Spike interactions with its human cellular receptor, ACE2. Here we show that changes in the inter-protomer energetics due to the D614G substitution favor a higher population of infection-capable (open) states. The inter-protomer interactions between S1 and S2 subunits in the open state of the D-form are asymmetric. This asymmetry is resolved in the G-form due to the release of tensile hydrogen bonds resulting in an increased population of open conformations. Thus, the increased infectivity of the G-form is likely due to a higher rate of profitable binding encounters with the host receptor. It is also predicted to be more neutralization sensitive due to enhanced exposure of the receptor binding domain, a key target region for neutralizing antibodies.

scholarly journals The SARS-CoV-2 Spike variant D614G favors an open conformational state

2021 ◽  
Vol 7 (16) ◽  
pp. eabf3671
Author(s):  
Rachael A. Mansbach ◽  
Srirupa Chakraborty ◽  
Kien Nguyen ◽  
David C. Montefiori ◽  
Bette Korber ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Molecular Level ◽  
Conformational State ◽  
Receptor Binding Domain ◽  
Upper Airways ◽  
Target Region ◽  
Viral Loads ◽  
Rapid Transition ◽  
Insight Into

The COVID-19 (coronavirus disease 2019) pandemic underwent a rapid transition with the emergence of a dominant viral variant (from the “D-form” to the “G-form”) that carried an amino acid substitution D614G in its “Spike” protein. The G-form is more infectious in vitro and is associated with increased viral loads in the upper airways. To gain insight into the molecular-level underpinnings of these characteristics, we used microsecond all-atom simulations. We show that changes in the protein energetics favor a higher population of infection-capable states in the G-form through release of asymmetry present in the D-form inter-protomer interactions. Thus, the increased infectivity of the G-form is likely due to a higher rate of profitable binding encounters with the host receptor. It is also predicted to be more neutralization sensitive owing to enhanced exposure of the receptor binding domain, a key target region for neutralizing antibodies. These results are critical for vaccine design.

scholarly journals Structural classification of neutralizing antibodies against the SARS-CoV-2 spike receptor-binding domain suggests vaccine and therapeutic strategies

2020 ◽  
Author(s):  
Christopher O. Barnes ◽  
Claudia A. Jette ◽  
Morgan E. Abernathy ◽  
Kim-Marie A. Dam ◽  
Shannon R. Esswein ◽  
...  
Keyword(s):  
Immune Responses ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Health Crisis ◽  
Naturally Occurring ◽  
Insight Into

AbstractThe COVID-19 pandemic presents an urgent health crisis. Human neutralizing antibodies (hNAbs) that target the host ACE2 receptor-binding domain (RBD) of the SARS-CoV-2 spike1–5 show therapeutic promise and are being evaluated clincally6–8. To determine structural correlates of SARS-CoV-2 neutralization, we solved 8 new structures of distinct COVID-19 hNAbs5 in complex with SARS-CoV-2 spike trimer or RBD. Structural comparisons allowed classification into categories: (1) VH3-53 hNAbs with short CDRH3s that block ACE2 and bind only to “up” RBDs, (2) ACE2-blocking hNAbs that bind both “up” and “down” RBDs and can contact adjacent RBDs, (3) hNAbs that bind outside the ACE2 site and recognize “up” and “down” RBDs, and (4) Previously-described antibodies that do not block ACE2 and bind only “up” RBDs9. Class 2 comprised four hNAbs whose epitopes bridged RBDs, including a VH3-53 hNAb that used a long CDRH3 with a hydrophobic tip to bridge between adjacent “down” RBDs, thereby locking spike into a closed conformation. Epitope/paratope mapping revealed few interactions with host-derived N-glycans and minor contributions of antibody somatic hypermutations to epitope contacts. Affinity measurements and mapping of naturally-occurring and in vitro-selected spike mutants in 3D provided insight into the potential for SARS-CoV-2 escape from antibodies elicited during infection or delivered therapeutically. These classifications and structural analyses provide rules for assigning current and future human RBD-targeting antibodies into classes, evaluating avidity effects, suggesting combinations for clinical use, and providing insight into immune responses against SARS-CoV-2.

scholarly journals Stereotypic neutralizing VH antibodies against SARS-CoV-2 spike protein receptor binding domain in COVID-19 patients and healthy individuals

2021 ◽  
pp. eabd6990
Author(s):  
Sang Il Kim ◽  
Jinsung Noh ◽  
Sujeong Kim ◽  
Younggeun Choi ◽  
Duck Kyun Yoo ◽  
...  
Keyword(s):  
B Cells ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
De Novo ◽  
Binding Domain ◽  
Host Cells ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Healthy Individuals

Stereotypic antibody clonotypes exist in healthy individuals and may provide protective immunity against viral infections by neutralization. We observed that 13 out of 17 patients with COVID-19 had stereotypic variable heavy chain (VH) antibody clonotypes directed against the receptor-binding domain (RBD) of SARS-CoV-2 spike protein. These antibody clonotypes were comprised of immunoglobulin heavy variable (IGHV)3-53 or IGHV3-66 and immunoglobulin heavy joining (IGHJ)6 genes. These clonotypes included IgM, IgG3, IgG1, IgA1, IgG2, and IgA2 subtypes and had minimal somatic mutations, which suggested swift class switching after SARS-CoV-2 infection. The different immunoglobulin heavy variable chains were paired with diverse light chains resulting in binding to the RBD of SARS-CoV-2 spike protein. Human antibodies specific for the RBD can neutralize SARS-CoV-2 by inhibiting entry into host cells. We observed that one of these stereotypic neutralizing antibodies could inhibit viral replication in vitro using a clinical isolate of SARS-CoV-2. We also found that these VH clonotypes existed in six out of 10 healthy individuals, with IgM isotypes predominating. These findings suggest that stereotypic clonotypes can develop de novo from naïve B cells and not from memory B cells established from prior exposure to similar viruses. The expeditious and stereotypic expansion of these clonotypes may have occurred in patients infected with SARS-CoV-2 because they were already present.

scholarly journals Murine monoclonal antibodies against RBD of SARS-CoV-2 neutralize authentic wild type SARS-CoV-2 as well as B.1.1.7 and B.1.351 viruses and protect in vivo in a mouse model in a neutralization dependent manner

2021 ◽  
Author(s):  
Fatima Amanat ◽  
Shirin Strohmeier ◽  
Wen-Hsin Lee ◽  
Sandhya Bangaru ◽  
Andrew B Ward ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Severe Acute Respiratory Syndrome ◽  
Mouse Model ◽  
Neutralizing Antibodies ◽  
Dependent Manner ◽  
Receptor Binding Domain ◽  
Wild Type ◽  
Murine Monoclonal Antibodies

After first emerging in December 2019 in China, severe acute respiratory syndrome 2 (SARS-CoV-2) has since caused a pandemic leading to millions of infections and deaths worldwide. Vaccines have been developed and authorized but supply of these vaccines is currently limited. With new variants of the virus now emerging and spreading globally, it is essential to develop therapeutics that are broadly protective and bind conserved epitopes in the receptor binding domain (RBD) or the whole spike of SARS-CoV-2. In this study, we have generated mouse monoclonal antibodies (mAbs) against different epitopes on the RBD and assessed binding and neutralization against authentic SARS-CoV-2. We have demonstrated that antibodies with neutralizing activity, but not non-neutralizing antibodies, lower viral titers in the lungs when administered in a prophylactic setting in vivo in a mouse challenge model. In addition, most of the mAbs cross-neutralize the B.1.351 as well as the B.1.1.7 variants in vitro.

scholarly journals SARS-CoV-2 escape in vitro from a highly neutralizing COVID-19 convalescent plasma

2020 ◽  
Author(s):  
Emanuele Andreano ◽  
Giulia Piccini ◽  
Danilo Licastro ◽  
Lorenzo Casalino ◽  
Nicole V. Johnson ◽  
...  
Keyword(s):  
South Africa ◽  
Immune Response ◽  
Antibody Response ◽  
Neutralizing Antibodies ◽  
Receptor Binding Domain ◽  
The United Kingdom ◽  
Effective Immune Response ◽  
Recent Emergence ◽  
Natural Variants

ABSTRACTTo investigate the evolution of SARS-CoV-2 in the immune population, we co-incubated authentic virus with a highly neutralizing plasma from a COVID-19 convalescent patient. The plasma fully neutralized the virus for 7 passages, but after 45 days, the deletion of F140 in the spike N-terminal domain (NTD) N3 loop led to partial breakthrough. At day 73, an E484K substitution in the receptor-binding domain (RBD) occurred, followed at day 80 by an insertion in the NTD N5 loop containing a new glycan sequon, which generated a variant completely resistant to plasma neutralization. Computational modeling predicts that the deletion and insertion in loops N3 and N5 prevent binding of neutralizing antibodies. The recent emergence in the United Kingdom and South Africa of natural variants with similar changes suggests that SARS-CoV-2 has the potential to escape an effective immune response and that vaccines and antibodies able to control emerging variants should be developed.One Sentence SummaryThree mutations allowed SARS-CoV-2 to evade the polyclonal antibody response of a highly neutralizing COVID-19 convalescent plasma.

scholarly journals Potent Neutralizing Antibodies Directed to Multiple Epitopes on SARS-CoV-2 Spike

2020 ◽  
Author(s):  
Lihong Liu ◽  
Pengfei Wang ◽  
Manoj S. Nair ◽  
Jian Yu ◽  
Micah Rapp ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Neutralizing Antibodies ◽  
Global Economy ◽  
Epitope Mapping ◽  
Severe Disease ◽  
Clinical Development ◽  
Receptor Binding Domain ◽  
Cryo Electron Microscopy ◽  
Novel Coronavirus

AbstractThe SARS-CoV-2 pandemic rages on with devasting consequences on human lives and the global economy1,2. The discovery and development of virus-neutralizing monoclonal antibodies could be one approach to treat or prevent infection by this novel coronavirus. Here we report the isolation of 61 SARS-CoV-2-neutralizing monoclonal antibodies from 5 infected patients hospitalized with severe disease. Among these are 19 antibodies that potently neutralized the authentic SARS-CoV-2 in vitro, 9 of which exhibited exquisite potency, with 50% virus-inhibitory concentrations of 0.7 to 9 ng/mL. Epitope mapping showed this collection of 19 antibodies to be about equally divided between those directed to the receptor-binding domain (RBD) and those to the N-terminal domain (NTD), indicating that both of these regions at the top of the viral spike are immunogenic. In addition, two other powerful neutralizing antibodies recognized quaternary epitopes that are overlapping with the domains at the top of the spike. Cryo-electron microscopy reconstructions of one antibody targeting RBD, a second targeting NTD, and a third bridging two separate RBDs revealed recognition of the closed, “all RBD-down” conformation of the spike. Several of these monoclonal antibodies are promising candidates for clinical development as potential therapeutic and/or prophylactic agents against SARS-CoV-2.

scholarly journals Neutralizing antibodies targeting the SARS-CoV-2 receptor binding domain isolated from a naïve human antibody library

2021 ◽  
Author(s):  
Benjamin Nikola Bell ◽  
Abigail E. Powell ◽  
Carlos Rodriguez ◽  
Jennifer R Cochran ◽  
Peter S. Kim
Keyword(s):  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
In Vitro Selection ◽  
Surface Display ◽  
Binding Domain ◽  
Yeast Surface Display ◽  
Surface Glycoprotein ◽  
Human Antibody ◽  
Receptor Binding Domain

Infection with SARS-CoV-2 elicits robust antibody responses in some patients, with a majority of the response directed at the receptor binding domain (RBD) of the spike surface glycoprotein. Remarkably, many patient-derived antibodies that potently inhibit viral infection harbor few to no mutations from the germline, suggesting that naive antibody libraries are a viable means for discovery of novel SARS-CoV-2 neutralizing antibodies. Here, we used a yeast surface-display library of human naive antibodies to isolate and characterize three novel neutralizing antibodies that target the RBD: one that blocks interaction with angiotensin-converting enzyme 2 (ACE2), the human receptor for SARS-CoV-2, and two that target other epitopes on the RBD. These three antibodies neutralized SARS-CoV-2 spike-pseudotyped lentivirus with IC50 values as low as 60 ng/mL in vitro. Using a biolayer interferometry-based binding competition assay, we determined that these antibodies have distinct but overlapping epitopes with antibodies elicited during natural COVID-19 infection. Taken together, these analyses highlight how in vitro selection of naive antibodies can mimic the humoral response in vivo, yielding neutralizing antibodies and various epitopes that can be effectively targeted on the SARS-CoV-2 RBD.

scholarly journals Thyroid Peroxidase Activity is Inhibited by Phenolic Compounds—Impact of Interaction

Molecules ◽  
2019 ◽  
Vol 24 (15) ◽  
pp. 2766
Author(s):  
Ewa Habza-Kowalska ◽  
Agnieszka A. Kaczor ◽  
Justyna Żuk ◽  
Dariusz Matosiuk ◽  
Urszula Gawlik-Dziki
Keyword(s):  
Chlorogenic Acid ◽  
Rosmarinic Acid ◽  
Molecular Level ◽  
Combination Index ◽  
3D Structure ◽  
Thyroid Peroxidase ◽  
Ic50 Values ◽  
Insight Into ◽  
Rutin And Quercetin

The aim of this study was to estimate the mode of thyroid peroxidase (TPO) inhibition by polyphenols: Chlorogenic acid, rosmarinic acid, quercetin, and rutin. All the tested polyphenols inhibited TPO; the IC50 values ranged from 0.004 mM to 1.44 mM (for rosmarinic acid and rutin, respectively). All these pure phytochemical substances exhibited different modes of TPO inhibition. Rutin and rosmarinic acid showed competitive, quercetin—uncompetitive and chlorogenic acid—noncompetitive inhibition effect on TPO. Homology modeling was used to gain insight into the 3D structure of TPO and molecular docking was applied to study the interactions of the inhibitors with their target at the molecular level. Moreover, the type and strength of mutual interactions between the inhibitors (expressed as the combination index, CI) were analyzed. Slight synergism, antagonism, and moderate antagonism were found in the case of the combined addition of the pure polyphenols. Rutin and quercetin as well as rutin and rosmarinic acid acted additively (CI = 0.096 and 1.06, respectively), while rutin and chlorogenic acid demonstrated slight synergism (CI = 0.88) and rosmarinic acid with quercetin and rosmarinic acid with chlorogenic acid showed moderate antagonism (CI = 1.45 and 1.25, respectively). The mixture of chlorogenic acid and quercetin demonstrated antagonism (CI = 1.79). All the polyphenols showed in vitro antiradical ability against 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid), ABTS. The highest ability (expressed as IC50) was exhibited by rosmarinic acid (0.12 mM) and the lowest value was ascribed to quercetin (0.45 mM).

scholarly journals Tat-Based Therapies as an Adjuvant for an HIV-1 Functional Cure

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 415 ◽  
Author(s):  
Hongping Jin ◽  
Dongsheng Li ◽  
Min-Hsuan Lin ◽  
Li Li ◽  
David Harrich
Keyword(s):  
Neutralizing Antibodies ◽  
Critical Role ◽  
Functional Cure ◽  
Infected People ◽  
Viral Loads ◽  
Cellular Factors ◽  
Combined Antiretroviral Therapy ◽  
Recent Developments ◽  
Direct Binding ◽  
Transactivator Of Transcription

The human immunodeficiency virus type 1 (HIV) establishes a chronic infection that can be well controlled, but not cured, by combined antiretroviral therapy (cART). Interventions have been explored to accomplish a functional cure, meaning that a patient remains infected but HIV is undetectable in the blood, with the aim of allowing patients to live without cART. Tat, the viral transactivator of transcription protein, plays a critical role in controlling HIV transcription, latency, and viral rebound following the interruption of cART treatment. Therefore, a logical approach for controlling HIV would be to block Tat. Tackling Tat with inhibitors has been a difficult task, but some recent discoveries hold promise. Two anti-HIV proteins, Nullbasic (a mutant of Tat) and HT1 (a fusion of HEXIM1 and Tat functional domains) inhibit viral transcription by interfering with the interaction of Tat and cellular factors. Two small molecules, didehydro-cortistatin A (dCA) and triptolide, inhibit Tat by different mechanisms: dCA through direct binding and triptolide through enhanced proteasomal degradation. Finally, two Tat-based vaccines under development elicit Tat-neutralizing antibodies. These vaccines have increased the levels of CD4+ cells and reduced viral loads in HIV-infected people, suggesting that the new vaccines are therapeutic. This review summarizes recent developments of anti-Tat agents and how they could contribute to a functional cure for HIV.

scholarly journals A Recombinant Protein SARS-CoV-2 Candidate Vaccine Elicits High-titer Neutralizing Antibodies in Macaques.

2021 ◽  
Author(s):  
Gary Baisa ◽  
David Rancour ◽  
Keith Mansfield ◽  
Monika Burns ◽  
Lori Martin ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
High Titer ◽  
Binding Domain ◽  
Pseudotyped Virus ◽  
Receptor Binding Domain ◽  
Binding Assays ◽  
Neutralizing Activity ◽  
Terminal Domain

Abstract BackgroundVaccines that generate robust and long-lived protective immunity against SARS-CoV-2 infection are urgently required. MethodsWe assessed the potential of vaccine candidates based on the SARS-CoV-2 spike in cynomolgus macaques (M. fascicularis) by examining their ability to generate spike binding antibodies with neutralizing activity. Antigens were derived from two distinct regions of the spike S1 subunit, either the N-terminal domain or an extended C-terminal domain containing the receptor-binding domain and were fused to the human IgG1 Fc domain. Three groups of 2 animals each were immunized with either antigen, alone or in combination. The development of antibody responses was evaluated through 20 weeks post-immunization. ResultsA robust IgG response to the spike protein was detected as early as 2 weeks after immunization with either protein and maintained for over 20 weeks. Sera from animals immunized with antigens derived from the RBD were able to prevent binding of soluble spike proteins to the ACE2 receptor, shown by in vitro binding assays, while sera from animals immunized with the N-terminal domain alone lacked this activity. Crucially, sera from animals immunized with the extended receptor binding domain but not the N-terminal domain had potent neutralizing activity against SARS-CoV-2 pseudotyped virus, with titers in excess of 10,000, greatly exceeding that typically found in convalescent humans. Neutralizing activity persisted for more than 20 weeks. ConclusionsThese data support the utility of spike subunit-based antigens as a vaccine for use in humans.

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