scholarly journals A human monoclonal antibody potently pan-neutralizes SARS-CoV-2 VOCs by targeting RBD invariant sites

2021 ◽  
Author(s):  
Xiaofei Wang ◽  
Ao Hu ◽  
Xiangyu Chen ◽  
Yixin Zhang ◽  
Fei Yu ◽  
...  
Keyword(s):  
Rational Design ◽  
Virus Disease ◽  
Human Monoclonal Antibody ◽  
Receptor Binding Domain ◽  
Wild Type ◽  
Cryo Electron Microscopy ◽  
Global Pandemic ◽  
Binding Epitope

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic of novel corona virus disease (COVID-19). The neutralizing monoclonal antibodies (mAbs) targeting the receptor binding domain (RBD) of SARS-CoV-2 are among the most promising strategies to prevent and treat COVID-19. However, SARS-CoV-2 variants of concern (VOCs) profoundly reduced the efficacies of most of mAbs and vaccines approved for clinical use. Herein, we demonstrated mAb 35B5 efficiently neutralizes both wild-type (WT) SARS-CoV-2 and VOCs, including B.1.617.2 (delta) variant, in vitro and in vivo. Cryo-electron microscopy (cryo-EM) revealed that 35B5 neutralizes SARS-CoV-2 by targeting a unique epitope that avoids the prevailing mutation sites on RBD identified in circulating VOCs, providing the molecular basis for its pan-neutralizing efficacy. The 35B5-binding epitope could also be exploited for the rational design of a universal SARS-CoV-2 vaccine.

scholarly journals Production of Noncapped Genomic RNAs Is Critical to Sindbis Virus Disease and Pathogenicity

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
Author(s):  
Autumn T. LaPointe ◽  
V Douglas Landers ◽  
Claire E. Westcott ◽  
Kevin J. Sokoloski
Keyword(s):  
Sindbis Virus ◽  
Virus Disease ◽  
Severe Disease ◽  
Wild Type Virus ◽  
Wild Type ◽  
Genomic Rnas ◽  
Mortality And Morbidity ◽  
The Impact

ABSTRACT Alphaviruses are positive-sense RNA viruses that utilize a 5′ cap structure to facilitate translation of viral proteins and to protect the viral RNA genome. Nonetheless, significant quantities of viral genomic RNAs that lack a canonical 5′ cap structure are produced during alphaviral replication and packaged into viral particles. However, the role/impact of the noncapped genomic RNA (ncgRNA) during alphaviral infection in vivo has yet to be characterized. To determine the importance of the ncgRNA in vivo, the previously described D355A and N376A nsP1 mutations, which increase or decrease nsP1 capping activity, respectively, were incorporated into the neurovirulent AR86 strain of Sindbis virus to enable characterization of the impact of altered capping efficiency in a murine model of infection. Mice infected with the N376A nsP1 mutant exhibited slightly decreased rates of mortality and delayed weight loss and neurological symptoms, although levels of inflammation in the brain were similar to those of wild-type infection. Although the D355A mutation resulted in decreased antiviral gene expression and increased resistance to interferon in vitro, mice infected with the D355A mutant showed significantly reduced mortality and morbidity compared to mice infected with wild-type virus. Interestingly, expression of proinflammatory cytokines was found to be significantly decreased in mice infected with the D355A mutant, suggesting that capping efficiency and the production of ncgRNA are vital to eliciting pathogenic levels of inflammation. Collectively, these data indicate that the ncgRNA have important roles during alphaviral infection and suggest a novel mechanism by which noncapped viral RNAs aid in viral pathogenesis. IMPORTANCE Mosquito-transmitted alphaviruses have been the cause of widespread outbreaks of disease that can range from mild illness to lethal encephalitis or severe polyarthritis. There are currently no safe and effective vaccines or therapeutics with which to prevent or treat alphaviral disease, highlighting the need to better understand alphaviral pathogenesis to develop novel antiviral strategies. This report reveals production of noncapped genomic RNAs (ncgRNAs) to be a novel determinant of alphaviral virulence and offers insight into the importance of inflammation to pathogenesis. Taken together, the findings reported here suggest that the ncgRNAs contribute to alphaviral pathogenesis through the sensing of the ncgRNAs during alphaviral infection and are necessary for the development of severe disease.

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 Natural Compounds from Djiboutian Medicinal Plants as Inhibitors of COVID-19 by in Silico Investigations

2020 ◽  
Author(s):  
abdirahman elmi ◽  
S. al jawad sayem ◽  
Mohamed Ahmed ◽  
fatouma mohamed
Keyword(s):  
Medicinal Plants ◽  
Natural Compounds ◽  
Binding Energies ◽  
Receptor Binding Domain ◽  
Global Pandemic ◽  
Main Protease ◽  
Target Sites ◽  
Better Than

The new coronavirus type SARS-Cov 2 (severe acute respiratory syndrome), which appeared in autumn 2019 in China, became a global pandemic in a few months. In this work, we looked for the potential anti SARS-Cov 2 of the compounds isolated from three Djiboutian medicinal plants namely Acacia seyal, Cymbopogon commutatus, and Indigofera caerulea. For this we carried out a docking with nine biomolecules, β-Sitosterol , Quercetin, Catechin, Lupeol, Rutin, Kaempferol, Gallic acid, Piperitone and Limonene on three target sites which are SARS-CoV-2 main protease (Mp), SARS-CoV-2 receptor binding domain (RBD) and human furin protease. These targets are chosen because of their role in the process of penetration of the virus into human cells and its multiplication. The phenolic compounds have a very good afinity on these three target sites with binding energies of up to -9.098 kcal/mol for rutin on SARS-CoV-2 Mp, much better than the two reference drugs hydroxychloroquine (-5.816 kcal / mol) and remdesivir (-7.194 kcal/mol). These natural compounds do not present toxicities and can be used pending In vitro and In vivo evaluations.

scholarly journals Reduced Infectivity in Cattle for an Outer Membrane Protein Mutant of Anaplasma marginale

2015 ◽  
Vol 81 (6) ◽  
pp. 2206-2214 ◽  
Author(s):  
Francy L. Crosby ◽  
Kelly A. Brayton ◽  
Forgivemore Magunda ◽  
Ulrike G. Munderloh ◽  
Karen L. Kelley ◽  
...  
Keyword(s):  
Rational Design ◽  
Anaplasma Marginale ◽  
Transposon Mutagenesis ◽  
Developmental Cycle ◽  
Wild Type ◽  
Content Type ◽  
Tick Infection ◽  
Tick Transmission

ABSTRACTAnaplasma marginaleis the causative agent of anaplasmosis in cattle. Transposon mutagenesis of this pathogen using theHimar1system resulted in the isolation of anomp10operon insertional mutant referred to as theomp10::himar1mutant. The work presented here evaluated if this mutant had morphological and/or growth rate defects compared to wild-typeA. marginale. Results showed that the morphology, developmental cycle, and growth in tick and mammalian cell cultures are similar for the mutant and the wild type. Tick transmission experiments established that tick infection levels with the mutant were similar to those with wild-typeA. marginaleand that infected ticks successfully infected cattle. However, this mutant exhibited reduced infectivity and growth in cattle. The possibility of transformingA. marginaleby transposon mutagenesis coupled within vitroandin vivoassessment of altered phenotypes can aid in the identification of genes associated with virulence. The isolation of deliberately attenuated organisms that can be evaluated in their natural biological system is an important advance for the rational design of vaccines against this species.

scholarly journals The functions of SARS-CoV-2 neutralizing and infection-enhancing antibodies in vitro and in mice and nonhuman primates

2021 ◽  
Author(s):  
Dapeng Li ◽  
Robert J Edwards ◽  
Kartik Manne ◽  
David R. Martinez ◽  
Alexandra Schäfer ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Nonhuman Primates ◽  
Safety Concern ◽  
Lung Pathology ◽  
Receptor Binding Domain ◽  
Biologically Relevant ◽  
Cryo Electron Microscopy ◽  
History Of

SummarySARS-CoV-2 neutralizing antibodies (NAbs) protect against COVID-19, making them a focus of vaccine design. A safety concern regarding SARS-CoV-2 antibodies is whether they mediate disease enhancement. Here, we isolated potent NAbs against the receptor-binding domain (RBD) and the N-terminal domain (NTD) of SARS-CoV-2 spike protein from individuals with acute or convalescent SARS-CoV-2 or a history of SARS-CoV-1 infection. Cryo-electron microscopy of RBD and NTD antibodies demonstrated function-specific modes of antibody binding. Select RBD NAbs also demonstrated Fc receptor-γ (FcγR)-mediated enhancement of virus infection in vitro, while five non-neutralizing NTD antibodies mediated FcγR-independent in vitro infection enhancement. However, both in vitro neutralizing and infection-enhancing RBD or infection-enhancing NTD antibodies protected from SARS-CoV-2 challenge in non-human primates and mice. One of 30 monkeys infused with enhancing antibodies had lung pathology and bronchoalveolar lavage cytokine evidence suggestive of enhanced disease. Thus, these in vitro assessments of enhanced antibody-mediated infection do not necessarily indicate biologically relevant in vivo infection enhancement.

scholarly journals Broad ultra-potent neutralization of SARS-CoV-2 variants by monoclonal antibodies specific to the tip of RBD

2021 ◽  
Author(s):  
Hang Ma ◽  
Yingying Guo ◽  
Haoneng Tang ◽  
Chien-Te Tseng ◽  
Lei Wang ◽  
...  
Keyword(s):  
Hydrophobic Effect ◽  
Receptor Binding Domain ◽  
Contact Interface ◽  
Block Interaction ◽  
Cryo Electron Microscopy ◽  
Binding Epitope ◽  
Clinical Illness ◽  
Animal Safety ◽  
Potent Neutralization

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) continue to wreak havoc across the globe. Higher transmissibility and immunologic resistance of VOCs bring unprecedented challenges to epidemic extinguishment. Here we describe a monoclonal antibody, 2G1, that neutralizes all current VOCs and has surprising tolerance to mutations adjacent to or within its interaction epitope. Cryo-electron microscopy structure showed that 2G1 bound to the tip of receptor binding domain (RBD) of spike protein with small contact interface but strong hydrophobic effect, which resulted in nanomolar to sub-nanomolar affinities to spike proteins. The epitope of 2G1 on RBD partially overlaps with ACE2 interface, which gives 2G1 ability to block interaction between RBD and ACE2. The narrow binding epitope but high affinity bestow outstanding therapeutic efficacy upon 2G1 that neutralized VOCs with sub-nanomolar IC50 in vitro. In SARS-CoV-2 and Beta- and Delta- variant-challenged transgenic mice and rhesus macaque models, 2G1 protected animals from clinical illness and eliminated viral burden, without serious impact to animal safety. Mutagenesis experiments suggest that 2G1 could be potentially capable of dealing with emerging SARS-CoV-2 variants in future. This report characterized the therapeutic antibodies specific to the tip of spike against SARS-CoV-2 variants and highlights the potential clinical applications as well as for developing vaccine and cocktail therapy.

scholarly journals Mechanistic insights into global suppressors of protein folding defects

2021 ◽  
Author(s):  
Gopinath Chattopadhyay ◽  
Jayantika Bhowmick ◽  
Kavyashree Manjunath ◽  
Shahbaz Ahmed ◽  
Parveen Goyal ◽  
...  
Keyword(s):  
Bacterial Toxin ◽  
Amino Acid Substitutions ◽  
Receptor Binding Domain ◽  
Loss Of Function ◽  
Wild Type ◽  
Partial Loss ◽  
Suppressor Mutations ◽  
Key Drivers

Most amino acid substitutions in a protein either lead to partial loss of function or are near neutral. Several studies have shown the existence of second-site mutations that can rescue defects caused by diverse loss of function mutations. Such global suppressor mutations are key drivers of protein evolution. However, the mechanisms responsible for such suppression remain poorly understood. To address this, we characterized multiple suppressor mutations both in isolation and in combination with inactive mutants. We examined five global suppressors of the bacterial toxin CcdB, the known M182T global suppressor of TEM-1 β-lactamase, the N239Y global suppressor of p53-DBD and three suppressors of the SARS-CoV-2 spike Receptor Binding Domain. The suppressors both alone, and in conjunction with inactive mutants, stabilise the protein both thermodynamically and kinetically in-vitro, predominantly through acceleration of the refolding rate parameters. When coupled to inactive mutants they promote increased in-vivo solubilities as well as regain-of-function phenotypes. Our study also demonstrates that the global suppressor approach can be used to consistently stabilise wild-type proteins, including for downstream translational applications.

scholarly journals In vitro Functional Analysis of pgRNA Sites Regulating Assembly of Hepatitis B Virus

2021 ◽  
Author(s):  
Nikesh Patel ◽  
Sam Clark ◽  
Eva U. Weiß ◽  
Carlos P. Mata ◽  
Jen Bohon ◽  
...  
Keyword(s):  
Symmetry Axis ◽  
Core Protein ◽  
Wild Type ◽  
Sequence Structure ◽  
Stem Loop ◽  
X Ray ◽  
Cryo Electron Microscopy ◽  
B Virus

AbstractThe roles of RNA sequence/structure motifs, Packaging Signals (PSs), for regulating assembly of an HBV genome transcript have been investigated in an efficient in vitro assay containing only core protein (Cp) and RNA. Variants of three conserved PSs, within the genome of a strain not used previously, preventing correct presentation of a Cp-recognition loop motif are differentially deleterious for assembly of nucleocapsid-like particles (NCPs). Cryo-electron microscopy reconstruction of the T=4 NCPs formed with the wild-type gRNA transcript, reveal that the interior of the Cp shell is in contact with lower resolution density, potentially encompassing the arginine-rich protein domains and gRNA. Symmetry-relaxation of this reconstruction reveals that such contacts are made at every symmetry axis. We infer from their regulation of assembly that some of these contacts would involve gRNA PSs, and confirmed this by X-ray RNA footprinting. Mutation of the ε stem-loop in the gRNA, where polymerase binds in vivo, produces a poor RNA assembly substrate with Cp alone, largely due to alterations in its conformation. The results show that RNA PSs regulate assembly of HBV genomic transcripts in vitro, and therefore may play similar roles in vivo, in concert with other molecular factors.

scholarly journals CONCEPTUAL REVIEW ON THE POTENTIAL OF PUNARNAVADYA GHRITA IN TACKLING THE SYMPTOMS OF COVID-19

2021 ◽  
Vol 12 (3) ◽  
pp. 157-163
Author(s):  
G Siva Ram
Keyword(s):  
Cannabis Sativa ◽  
Virus Disease ◽  
World Health Organisation ◽  
World Health ◽  
Corona Virus ◽  
Global Pandemic ◽  
Phytochemical Compounds ◽  
Effective Remedy

COVID-19 disease has created panic among all with respect to health and economy. On 11th March 2020 WHO (world health organisation) declared novel corona virus outbreak as global pandemic. Confirmed effective remedy to prevent or treat COVID-19 is not yet established. Ayurveda, the Indian system of medicine is the oldest medicinal literature known to mankind as old as 3000 BC and is recognised as medical system by WHO. There is a need to investigate time tested potent classical Ayurvedic formulations backed by contemporary scientific studies in treating symptoms of COVID-19 disease. Vijaya (Cannabis sativa Linn.) in Ayurveda notable for its Vyavayi (quick absorption) and Yogavahi (synergetic action) properties said to be originated from ambrosia as per Vedic mythology. Recent scientific research on the phytochemicals of Cannabis plant showed promising results in decreasing the pro-inflammatory cytokine storm and to an extent halting the replication of SARS-CoV-2. We have put forward one such classical Cannabis Ayurvedic formulation Punarnavadya Ghrita available in market which can be useful as an alternative remedy for prophylactic and curative to the symptoms of novel corona virus disease whose individual herbal phytochemical compounds are studied through in silico, in vitro and in vivo methods in treating COVID-19 with positive outcomes.

scholarly journals Isolation and comparative analysis of antibodies that broadly neutralize sarbecoviruses

2021 ◽  
Author(s):  
Lihong Liu ◽  
Sho Iketani ◽  
Yicheng Guo ◽  
Ryan Casner ◽  
Eswar Reddem ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Comparative Analysis ◽  
Receptor Binding ◽  
Comparative Studies ◽  
Human Monoclonal Antibody ◽  
Receptor Binding Domain ◽  
Pandemic Preparedness ◽  
Antibody Class

The devastation caused by SARS-CoV-2 has made clear the importance of pandemic preparedness. To address future zoonotic outbreaks due to related viruses in the sarbecovirus subgenus, we identified a human monoclonal antibody, 10-40, that neutralized or bound all sarbecoviruses tested in vitro and protected against SARS-CoV-2 and SARS-CoV in vivo. Comparative studies with other receptor-binding domain (RBD)-directed antibodies showed 10-40 to have the greatest breadth against sarbecoviruses and thus its promise as an agent for pandemic preparedness. Moreover, structural analyses on 10-40 and similar antibodies not only defined an epitope cluster in the inner face of the RBD that is well conserved among sarbecoviruses, but also uncovered a new antibody class with a common CDRH3 motif. Our analyses also suggested that elicitation of this class of antibodies may not be overly difficult, an observation that bodes well for the development of a pan-sarbecovirus vaccine.

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