MERS-CoV Spike protein (aa 1-1297) [His] (DAG-H10300)

MERS-CoV Spike protein (aa 1-1297) [His], recombinant protein from Baculovirus

Nature
Recombinant
Predicted N terminal
Tyr 18
Tag/Conjugate
His
Endotoxin
< 1.0 EU per μg of the protein as determined by the LAL method
Application Notes
ELISA: 10 μg/mL
Procedure
None
Purity
> 85 % as determined by SDS-PAGE
Format
Lyophilized from sterile 20mM Tris, 500mM Nacl, pH 7.4, 10% glycerol
Size
20μg; 50μg
Preservative
None
Storage
Store it under sterile conditions at -70 °C. It is recommended that the protein be aliquoted for optimal storage. Avoid repeated freeze-thaw cycles.
Stability
Samples are stable for up to twelve months from date of receipt at -70°C
Introduction
The spike (S) glycoprotein of coronaviruses contains protrusions that will only bind to certain receptors on the host cell: they are essential for both host specificity and viral infectivity. The term 'peplomer' is typically used to refer to a grouping of heterologous proteins on the virus surface that function together. The spike (S) glycoprotein of coronaviruses is known to be essential in the binding of the virus to the host cell at the advent of the infection process. Most notable is severe acute respiratory syndrome (SARS). The severe acute respiratory syndrome-coronavirus (SARS-CoV) spike (S) glycoprotein alone can mediate the membrane fusion required for virus entry and cell fusion. It is also a major immunogen and a target for entry inhibitors. The SARS-CoV spike (S) protein is composed of two subunits; the S1 subunit contains a receptor-binding domain that engages with the host cell receptor angiotensin-converting enzyme 2 and the S2 subunit mediates fusion between the viral and host cell membranes. The S protein plays key parts in the induction of neutralizing-antibody and T-cell responses, as well as protective immunity, during infection with SARS-CoV.
Keywords
Coronavirus; Corona; Coronaviridae; Coronavirinae

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References


Unraveling the Epidemiology, Geographical Distribution, and Genomic Evolution of Potentially Lethal Coronaviruses (SARS, MERS, and SARS CoV-2)

FRONTIERS IN CELLULAR AND INFECTION MICROBIOLOGY

Authors: Masood, Nosheen; Malik, Saima Shakil; Raja, Muhammad Naqqi; Mubarik, Sumaira; Yu, Chuanhua

SARS CoV appeared in 2003 in China, transmitted from bats to humans via eating infected animals. It affected 8,096 humans with a death rate of 11% affecting 21 countries. The receptor binding domain (RBD) in S protein of this virus gets attached with the ACE2 receptors present on human cells. MERS CoV was first reported in 2012 in Middle East, originated from bat and transmitted to humans through camels. MERS CoV has a fatality rate of 35% and last case reported was in 2017 making a total of 1,879 cases worldwide. DPP4 expressed on human cells is the main attaching site for RBD in S protein of MERS CoV. Folding of RBD plays a crucial role in its pathogenesis. Virus causing COVID-19 was named as SARS CoV-2 due its homology with SARS CoV that emerged in 2003. It has become a pandemic affecting nearly 200 countries in just 3 months' time with a death rate of 2-3% currently. The new virus is fast spreading, but it utilizes the same RBD and ACE2 receptors along with furin present in human cells. The lessons learned from the SARS and MERS epidemics are the best social weapons to face and fight against this novel global threat.

Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen

VACCINE

Authors: Nyon, Mun Peak; Du, Lanying; Tseng, Chien-Te Kent; Seid, Christopher A.; Pollet, Jeroen; Naceanceno, Kevin S.; Agrawal, Anurodh; Algaissi, Abdullah; Peng, Bi-Hung; Tai, Wanbo; Jiang, Shibo; Bottazzi, Maria Elena; Strych, Ulrich; Hotez, Peter J.

Middle East respiratory syndrome coronavirus (MERS-CoV) has infected at least 2040 patients and caused 712 deaths since its first appearance in 2012, yet neither pathogen-specific therapeutics nor approved vaccines are available. To address this need, we are developing a subunit recombinant protein vaccine comprising residues 377-588 of the MERS-CoV spike protein receptor-binding domain (RBD), which, when formulated with the AddaVax adjuvant, it induces a significant neutralizing antibody response and protection against MERS-CoV challenge in vaccinated animals. To prepare for the manufacture and first-in-human testing of the vaccine, we have developed a process to stably produce the recombinant MERS S377-588 protein in Chinese hamster ovary (CHO) cells. To accomplish this, we transfected an adherent dihydrofolate reductase-deficient CHO cell line (adCHO) with a plasmid encoding S377-588 fused with the human IgG Fc fragment (S377-588-Fc). We then demonstrated the interleukin-2 signal peptide-directed secretion of the recombinant protein into extracellular milieu. Using a gradually increasing methotrexate (MTX) concentration to 5 mu M, we increased protein yield by a factor of 40. The adCHO-expressed S377-588-Fc recombinant protein demonstrated functionality and binding specificity identical to those of the protein from transiently transfected HEK293T cells. In addition, hCD26/dipeptidyl peptidase-4 (DPP4) transgenic mice vaccinated with AddaVax-adjuvanted S377-588-Fc could produce neutralizing antibodies against MERS-CoV and survived for at least 21 days after challenge with live MERS-CoV with no evidence of immunological toxicity or eosinophilic immune enhancement. To prepare for large scale-manufacture of the vaccine antigen, we have further developed a high-yield monoclonal suspension CHO cell line. (C) 2018 Elsevier Ltd. All rights reserved.

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