Dynamic remodeling of USP28 by the selective inhibitor CAS-010: Insights from DFT and molecular dynamics simulations

Area of Science:

• Biochemistry
• Structural Biology
• Computational Chemistry

Background:

• Ubiquitin-specific protease 28 (USP28) is a deubiquitinase crucial for cancer progression by stabilizing oncoproteins like c-Myc.
• USP28 is a significant anti-cancer target due to its role in tumorigenesis.
• The inhibitor CAS-010 shows high selectivity and potency against USP28, but its binding mechanism is not fully understood.

Purpose of the Study:

• To elucidate the dynamic binding mechanism of CAS-010 to USP28.
• To investigate how CAS-010 binding affects USP28 conformation and catalytic function.
• To provide a structural basis for the rational design of novel USP28 inhibitors.

Main Methods:

• Density Functional Theory (DFT) calculations to assess inhibitor properties.
• Molecular Dynamics (MD) simulations (200 ns) to observe dynamic binding events.
• Principal Component Analysis (PCA), Dynamic Cross-Correlation Matrix (DCCM), and free energy landscape analyses to characterize conformational changes.
• Binding free energy calculations and residue-level decomposition (ASIE) to identify binding hotspots.

Main Results:

• DFT calculations indicated CAS-010's metabolic stability and binding compatibility.
• MD simulations revealed a biphasic dynamic remodeling: localized tightening of the catalytic pocket and distal allosteric relaxation.
• This remodeling effectively inactivates USP28 by restricting active-site flexibility.
• Strong spontaneous binding (ΔGbind ≈ -44 to -47 kcal/mol) was confirmed, with key anchoring residues identified (Phe370, Tyr643, His592, His261).

Conclusions:

• CAS-010 achieves potent USP28 inhibition through orchestrated dynamic remodeling, not global rigidification.
• The study provides a detailed mechanistic understanding of CAS-010's action on USP28.
• Findings offer valuable structural insights for developing next-generation USP28-targeted anti-cancer therapies.