Allostery in homodimeric SARS-CoV-2 main protease.

Area of Science:

• Biochemistry
• Structural Biology
• Virology

Background:

• Many enzymes function as homodimers with separate catalytic sites, but the functional advantage is often unclear.
• The SARS-CoV-2 main protease (Mpro) is crucial for viral replication, requiring dimerization for its activity and regulatory role.

Purpose of the Study:

• To investigate the allosteric mechanism of SARS-CoV-2 Mpro during substrate interaction.
• To understand how Mpro's plasticity accommodates different peptide substrates and mediates communication between active sites.

Main Methods:

• X-ray crystallography
• Native mass spectrometry
• Isothermal titration calorimetry
• Enzyme activity assays

Main Results:

• Mpro dimers exhibit asymmetric structures upon binding peptide substrates, unlike their free form.
• Allosteric communication between active sites was observed when substrates (nsp5/6, nsp14/15) bound to a single subunit.
• Arginines 4 and 298 were identified as critical residues for the symmetric-to-asymmetric dimer transition.
• Positive cooperativity was demonstrated through increased processing efficiency (kinetic allostery), not enhanced substrate binding (thermodynamic allostery).

Conclusions:

• SARS-CoV-2 Mpro employs kinetic allostery to regulate viral polyprotein processing.
• This mechanism allows for spatiotemporal control of viral replication at the physiological level.
• The plasticity of Mpro and specific residue interactions are key to its allosteric function.