3CL Protease, Untagged (SARS-CoV-2) Assay Kit

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78042
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Description

The Untagged 3CL Protease Assay Kit is designed to measure 3CL Protease activity for screening and profiling applications, in a homogeneous assay with no time-consuming washing steps. The kit comes in a convenient 96-well format, with purified untagged 3CL Protease (BPS Bioscience #100823), fluorogenic substrate, and 3CL Protease assay buffer for 100 enzyme reactions. 3CL inhibitor GC376 is also included as a control.

The 3CL Protease Substrate is an internally quenched 14-mer fluorogenic peptide (DABCYL-KTSAVLQSGFRKME-EDANS). When the donor (EDANS) and acceptor (DABCYL) fluorophores are in close proximity, the energy emitted from EDANS is quenched by DABCYL (intact substrate). Upon proteolysis by 3CL, the peptide substrate is cleaved between glutamine and serine by the 3CL protease to generate the highly fluorescent peptide fragment (SGFRKME-EDANS). EDANS has an excitation peak at 336 nm and an emission peak at 455 nm. The fluorescence intensity increases proportionally to the activity of 3CL. More information on the substrate, including MW and structure, can be found on our website (BPS Bioscience #79952).

Figure 1: Illustration of the principle behind the 3CL protease assay.
The 3CL Protease Substrate is an internally quenched 14-mer fluorogenic peptide (DABCYL-KTSAVLQSGFRKME-EDANS). When the donor (EDANS) and acceptor (DABCYL) fluorophores are in close proximity the energy emitted from EDANS is quenched by DABCYL (intact substrate). Upon proteolysis by 3CL, the peptide substrate is cleaved between the glutamine and serine residues to generate the highly fluorescent peptide fragment (SGFRKME-EDANS). The fluorescence intensity increases proportionally to the activity of 3CL More information on the substrate, including MW and structure, can be found on our website (BPS Bioscience #79952).

Assay Principle

3CL Protease Assay Kit

Need us to run inhibitor screens or profile your compounds against 3CL Protease, Untagged (SARS-CoV-2)? Check out our Protease Screening Services or Coronavirus Services

This product has been cited 39 times.

Synonyms
main protease kit, Mpro, SARS-CoV-2 3CL Protease kit, untagged 3CL kit, 3 CL assay, 3CL Pro kit, nsp5
Product Info
Storage and Usage
Citations39
Assay Kit Format
Fluorogenic
Supplied As
The kit comes in a convenient 96-well format or 384-well format, with purified untagged 3CL Protease, fluorogenic substrate, and 3CL Protease assay buffer for 100 or 384 enzyme reactions . 3CL inhibitor GC376 is also included as a positive control.
Materials Required But Not Supplied

Fluorescent microplate reader capable of reading exc/em=360 nm/460 nm

Format

96 Reactions:

Catalog # Component Amount Storage
100823 3CL Protease (SARS-CoV-2), no tag* >2 µg -80°C Avoid
freeze/
thaw
cycles!
79952 3CL Protease Substrate
(10 mM)
25 µl -80°C
79956 3CL Protease Assay Buffer 25 ml -20°C
78013 GC376, MW=507.5 50 µg -20°C
  0.5 M DTT 200 µl -20°C
79685 Black, low binding microtiter plate 1 Room
Temp
 
  Plate sealing film 1  

*The exact concentration of protein is lot-specific and will be indicated on the tube containing the protein. Excess material is provided for ease of retrieval.

 

384 Reactions:

Catalog # Component Amount Storage
100823 3CL Protease (SARS-CoV-2), no tag 6 µg -80°C Avoid
freeze/
thaw
cycles!
79952 3CL Protease Substrate
(10 mM)
50 µl -80°C
79956 3CL Protease Assay Buffer 25 ml -20°C
78013 GC376, MW=507.5 2 x 50 µg 20°C
  0.5 M DTT 200 µl -20°C
79961 Black, 384-well plate 1 Room
Temp
 
  Plate sealing film 1    

*The exact concentration of protein is lot-specific and will be indicated on the tube containing the protein.

 

Background

Coronaviruses (CoVs) cause respiratory and intestinal infections in humans and animals. The 3CL protease, also known as Main Protease (Mpro), plays a vital role in processing the polyproteins that are translated from the viral RNA. Protease inhibitors that can block viral replication are promising potential drug candidates for the treatment of patients suffering from COVID-19 infection.

References

1. Jared S. Morse, et al., 2020 Chem.Bio.Chem. 21:730–738.
2. Zhang, L., et al. 2020, Science 368 (6489): 409-412.