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SARS-CoV / SARS-CoV-2 Main Protease Fluorogenic Substrate 2020/06/23
Peptides Institute Inc. is pleased to announce three types of substrates for the SARS-CoV main protease (SARS-CoV Mpro or 3CLpro) and SARS-CoV-2 main protease (SARS-CoV-2 Mpro). Protease activity can be measured by observing the increase in fluorescence by the main protease cleaving the substrate, useful in the research of inhibitors.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative virus of a new coronavirus infection (COVID-19), which has been spreading worldwide since the end of 2019. When a virus infects a host cell, the RNA from the virus translates a large precursor protein. The precursor protein is cleaved by various proteases, and becomes a functional protein required for virus growth such as RNA-dependent RNA polymerase. SARS-CoV-2 Mpro, which is also one of the cleavage products, plays a major role in cleavage of precursor proteins and is an essential protease for viral growth. Therefore, this “main protease” inhibitor may be an antiviral drug and is being developed all over the world.
Dabcyl-Lys-Thr-Ser-Ala-Val-Leu-Gln-Ser-Gly-Phe-Arg-Lys-Met-Glu(Edans)-NH2
Fluorogenic substrate using FRET of SARS-CoV / SARS-CoV-2 Mpro1),2),3)
The precursor protein sequence, Thr-Ser-Ala-Val-Leu-Gln ↓ Ser-Gly-Phe-Arg-Lys-Met [↓ is a cleavage site] and a fluorescent group (Edans) and a quenching group (Dabcyl) are introduced. SARS-CoV Mpro cleaves the Gln-Ser sequence between the fluorescent and quencher groups, resulting in increased fluorescence. By observing this fluorescence, the protease activity can be measured [KM=17μM, kcat=1.9s-1 (kinetic parameter for SARS-CoV Mpro)]1) [KM=28μM (SARS-CoV-2 Kinetic parameters for Mpro)].3)
Figure 1. Method of measuring protease activity of SARS-CoV/SARS-CoV-2 Mpro quenching fluorogenic substrate using FRET.
| Code | Product Name | Quant |
| 3249-v | Dabcyl-Lys-Thr-Ser-Ala-Val-Leu-Gln-Ser-Gly-Phe-Arg-Lys-Met-Glu(Edans)-NH2 | 1 mg |
Ac-Abu-Tle-Leu-Gln-MCA・Ac-Thz-Tle-Leu-Gln-MCA
Fluorogenic substrate for SARS-CoV / SARS-CoV-2 Mpro 4)
4-methylcoumarin-7-amide (MCA) type fluorogenic substrate in which aminomethylcoumarin (AMC) is introduced into two sequences with high affinity for SARS-CoV-2 Mpro, as reported by Drag et al. When the non-fluorescent MCA-bound substrate is cleaved by SARS-CoV-2 Mpro, AMC with strong fluorescence is released. By observing this fluorescence, protease activity can be measured. (Ex: 380nm / Em: 460nm)
| Code | Product Name | Quant |
| 3250-v | Ac-Abu-Tle-Leu-Gln-MCA | 1 mg |
| 3251-v | Ac-Thz-Tle-Leu-Gln-MCA | 1 mg |
Other CoV related products
| Code | Product Name | Quant |
| 3252-v | Z-Arg-Leu-Arg-Gly-Gly-MCA (Substrate for PLPro) | 1 mg |
| 3206-v | Ubiquitin-MCA (Substrate for PLPro) | 50 μg |
| 3155-v | Boc-Arg-Val-Arg-Arg-MCA (Substrate for Furin) | 5 mg |
| 3159-v | Pyr-Arg-Thr-Lys-Arg-MCA (Substrate for Furin) | 5 mg |
| 3099-v | AMC (Reference Compound) | 5 mg |
| 3253-v | Ac-Abu-D-Tyr-Leu-Gln-VS (Inhibitor for Mpro) | 1 mg |
| 4041 | Leupeptin (Inhibitor for Serine Protease) | 25 mg |
| 4321-v | E-64-d (Inhibitor for Thiol Protease) | 5 mg |
- C.-J. Kuo, Y.-H. Chi, J.T.-A. Hsu, and P.-H. Liang, Biochem. Biophys. Res. Commun., 318, 862 (2004).
- L. Zhang, D. Lin, X. Sun, U. Curth, C. Drosten, L. Sauerhering, S. Becker, K. Rox, and R. Hilgenfeld, Science, 368, 409 (2020).
- M. D. Sacco, C. Ma, P. Lagarias, A. Gao, J. A. Townsend, X. Meng, P. Dube, X. Zhang, Y. Hu, N. Kitamura, B. Hurst, B. Tarbet, M. T. Marty, A. Kolocouris, Y. Xiang, Y. Chen, and J. Wang, Sci Adv., 6, eabe0751 (2020).
- W. Rut, K. Groborz, L. Zhang, X. Sun, M. Zmudzinski, B. Pawlik, X. Wang, D. Jochmans, J. Neyts, W. Młynarski, R. Hilgenfeld, and M. Drag, Nat. Chem. Biol., 17, 222 (2021).
