Potential blocker of SARS-CoV entry and a narrow functionality of its spike protein motifs on Qubevirus platform

Area of Science:

• Virology and Molecular Biology
• Immunology
• Biotechnology

Background:

• Targeting severe acute respiratory syndrome coronavirus (SARS-CoV) entry is crucial for antiviral development.
• The SARS-CoV spike (S) protein interacts with human angiotensin-converting enzyme 2 (hACE2) to mediate host cell entry.

Purpose of the Study:

• To identify the smallest active spike protein binding motif (RBSM) and key residues responsible for SARS-CoV binding to hACE2.
• To validate the potential of engineered RBSM variants as SARS-CoV entry inhibitors.
• To optimize a chimeric protein construct for anti-SARS-CoV antibody detection.

Main Methods:

• Utilized the Qubevirus (Qβ) platform for phage display of spike protein binding motif variants.
• Assessed disruption of hACE2 recognition and SARS-CoV pseudovirus infectivity using engineered RBSM.
• Constructed and optimized a chimera of S protein epitopes fused to Qβ minor coat protein for antibody titration.

Main Results:

• Identified five critical residues (L472, N473, N479, D480, Y491) within the smallest active spike protein binding motif (S_473-492).
• Engineered RBSM variants, including QβRBSM1, effectively blocked hACE2 recognition and reduced SARS-CoV pseudovirus infectivity without cytotoxicity.
• Determined an optimal chimera configuration of three S protein epitopes for sensitive anti-S antibody titration.

Conclusions:

• The identified RBSM and key residues are druggable targets for developing SARS-CoV entry inhibitors.
• Engineered RBSM variants show promise as blockers of SARS-CoV entry.
• The optimized chimeric construct provides a foundation for developing SARS-CoV antibody detection biosensors and subunit vaccines.