Cell-Based Assays to Fight COVID-19

As the COVID-19 pandemic becomes increasingly severe, many of the world’s researchers have focused urgently on this single topic in unprecedented ways. ACROBiosystems has also been a major contributor in this collective effort against COVID-19.

The spike of SARS-CoV-2 is a highly glycosylated homologous trimer made up of two subunits: S1 and S2.1,2 Like SARS-CoV-1, SARS-CoV-2’s spike protein enables viral entry into a target cell by exploiting the Angiotensin-converting enzyme 2 (ACE2) receptor.1,2,3

TMPRSS2 - a serine protease - is also a part of this process, taking responsibility for priming of the spike protein. Here, the proteolytic cleavage triggered the shedding of the S1 subunit and the ACE2.3,4

Because of this shedding, a far weaker signal under flow cytometry analysis is observed when examining the spike protein or S1 protein binding with ACE2 on the surface of the cell. This differs from the robust signal obtained from protein based SPR or ELISA analysis.

ACROBiosystems has developed an effective flow cytometry assay that can examine the binding between S1 protein/S1 RBD protein and ACE2. This assay can be used to neutralize antibody screening.

Cat. No.: S1N-C5257

SARS-CoV-2 (COVID-19) S1 protein, Mouse IgG2a Fc Tag

Cell-Based Assays to Fight COVID-19

Image Credit: ACROBiosystems

Cat. No.: SPD-C5259

SARS-CoV-2 (COVID-19) S protein RBD, Mouse IgG2a Fc Tag

Cell-Based Assays to Fight COVID-19

Image Credit: ACROBiosystems

The transmembrane protease ADAM17 has also been reported as being involved in the infection of host cells.5.6 Binding of S protein to ACE2 has been found to activate ADAM17, catalyzing the ACE2 shedding and assisting the virus’ entry into the cell.6,7,8,9

In 2010, Haga et al. showed that SARS-CoV-1 spike protein has the potential to induce ACE2 shedding in the VERO E6 cell model. They also highlighted that TAPI-2, which is the inhibitor of ADAM17, is able to prevent ACE2 shedding.6,9 ACROBiosystems has repeated this assay with SARS-CoV-2 spike protein.

In this instance, a fluorescent peptide substrate was employed to detect ACE2 activity within cell culture supernatant. It was confirmed that the S1 protein of SARS-CoV-2 was able to induce the shedding of ACE2 in VERO E6 cells – see the figure below.

SARS-CoV-2 (COVID-19) S1 protein, mouse IgG2a Fc tag

Cat. No.: S1N-C5257

SARS-CoV-2 (COVID-19) S1 protein, his tag

Cat. No.: S1N-C52H3

Cell-Based Assays to Fight COVID-19

Image Credit: ACROBiosystems

References

  1. Structural basis for the recognition of SARS-CoV-2 by full-length human ACE2. Science 367, 1444–1448(2020).
  2. Cryo-EM Structure of the2019-nCoV Spike in the Prefusion Conformation. Science 367, 1260-1263(2020).
  3. SARS-CoV-2 Cell entry depends on ACE2 and TMPRSS2 and Is blocked by a clinically proven protease Inhibitor. Cell 181, 1–10(2020).
  4. Structural Basis of SARS-CoV-2Spike Protein Priming by TMPRSS2. https://doi.org/10.1101/2020.04.21.052639.
  5. Tumor necrosis Factor-a convertase(ADAM17) mediates regulated ectodomain shedding of the severe-acute respiratory syndrome-coronavirus (SARS-CoV) receptor, angiotensin-converting enzyme-2(ACE2). JBC 280,30113–30119(2005).
  6. Modulation of TNF-a-converting enzyme by the spike protein of SARS-CoV and ACE2 induces TNF-a production and facilitates viral entry. PNAS 105, 7809–7814(2008)
  7. Trilogy of ACE2: A peptidase inthe renin–angiotensin system, a SARS receptor, and a partner for amino acid transporters. Pharmacology & Therapeutics 128, 119–128(2010).
  8. TACE antagonists blocking ACE2 shedding caused by the spike protein of SARS-CoV are candidate antiviral compounds. Antiviral Research, 85, 551–555(2010).
  9. Differential Downregulation of ACE2 by the Spike Proteins of Severe Acute Respiratory Syndrome Coronavirus and Human CoronavirusNL63. VIROLOGY, 84, 1198–1205(2010).

About ACROBiosystems

ACROBiosystems is a cornerstone enterprise of the pharmaceutical and biotechnology industries. Their mission is to help overcome challenges with innovative tools and solutions from discovery to the clinic. They supply life science tools designed to be used in discovery research and scalable to the clinical phase and beyond. By consistently adapting to new regulatory challenges and guidelines, ACROBiosystems delivers solutions, whether it comes through recombinant proteins, antibodies, assay kits, GMP-grade reagents, or custom services. ACROBiosystems empower scientists and engineers dedicated towards innovation to simplify and accelerate the development of new, better, and more affordable medicine.


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Last updated: Jan 29, 2024 at 11:20 AM

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