Biological context
Validated COVID-19 drug target. Nirmatrelvir (the active ingredient of Paxlovid, approved 2021) is a peptidomimetic covalent inhibitor that mimics the tetrahedral transition state.
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Hydrolase / cysteine protease · Peptidase C30 (coronavirus main protease)
SARS-CoV-2 main protease (Mpro / 3CLpro)
EC EC 3.4.22.69PDB 6LU7SARS-CoV-2
Cysteine-histidine catalytic dyad. Cleaves the coronaviral polyprotein at 11 conserved Leu-Gln↓(Ser/Ala/Gly) sites — essential for viral replication.
Mechanistic insight
Mpro is functional only as a homodimer; the N-terminal finger of one protomer organises the oxyanion hole and S1 pocket of the other. The dimer interface is itself a potential drug target ("allosteric inhibition by dimer disruption").
cysteine proteasecoronavirusdrug targetcovalent inhibitorSARS-CoV-2
Catalytic mechanism
Type: Cysteine-histidine dyad — covalent acyl-enzyme (thioester) intermediate
Rate-limiting: Acylation (for most peptide substrates)
Overall reaction
Polyprotein-Leu-Gln↓(Ser/Ala/Gly)-Xaa + H2O → two peptides
01
Substrate recognition in S1–S4 pockets
Mpro recognises the Leu-Gln↓(Ser/Ala/Gly) motif. Gln P1 hydrogen-bonds to His163, Glu166 and Phe140 in the S1 subsite — the Gln specificity is absolute and is what distinguishes Mpro from host cysteine proteases. Hydrophobic P2 (usually Leu) inserts into a pocket lined by His41, Met49 and Tyr54.
A:145A:41A:163A:166A:140
Mpro is only catalytically active as a homodimer; the N-terminal residues (the 'N-finger') of each protomer organise the S1 pocket of the other.
Step 1 / 6
References (3)
- Jin Z et al. Structure of Mpro from SARS-CoV-2 and discovery of its inhibitors. Nature 582:289–293 (2020).
- Owen DR et al. An oral SARS-CoV-2 Mpro inhibitor clinical candidate for the treatment of COVID-19. Science 374:1586–1593 (2021).
- Anand K et al. Coronavirus main proteinase (3CLpro) structure: basis for design of anti-SARS drugs. Science 300:1763–1767 (2003).
Kinetic parameters
Steady-state kcat, KM and specificity constants from literature.
| Substrate | kcat (s⁻¹) | KM (mM) | kcat/KM (M⁻¹s⁻¹) | pH | T (°C) | Source |
|---|---|---|---|---|---|---|
| MCA-AVLQ↓SGFR-Lys(Dnp)-K (FRET) | 0.300 | 0.028 | 1.07 × 10⁴ | 7.3 | 25 | Zhang et al., Science 2020 |
| Ac-Abu-Tle-Leu-Gln-MCA | 0.0450 | 0.16 | 2.81 × 10² | 7.5 | 25 | Kneller et al., Nat. Commun. 2020 |
Mpro is a relatively slow cysteine protease (kcat on the order of 0.1 s⁻¹). It compensates with strong substrate specificity for the Leu-Gln motif. Dimerisation is essential — monomeric Mpro is catalytically dead, making the dimer interface an allosteric drug target.
Michaelis–Menten · steady-state simulator
Run Mpro in silico.
Integrate the reaction v = Vmax·[S] / (Km,app + [S]) in closed form with RK4, across any [S], [E], and an optional reversible inhibitor. Defaults are drawn from the curated kinetic record for this enzyme where available.
Progress curve · [S](t) and [P](t)30 s window
substrate [S]product [P]
Saturation · v vs. [S]Km,app = 0.0280 mM · Vmax,app = 3.00e-4 mM/s
Inhibitor
Curated kinetic record
Defaults seeded from MCA-AVLQ↓SGFR-Lys(Dnp)-K (FRET) at pH 7.3 / 25 °C — source: Zhang et al., Science 2020.
Why this enzyme is in the catalog
Contemporary example of: cysteine protease mechanism, rapid structure-based drug discovery under pandemic conditions, covalent inhibitor design, pan-coronavirus scaffolds.