Structure-based design of prefusion-stabilized SARS-CoV-2 spikes

Area of Science:

• Biochemistry
• Structural Biology
• Virology

Background:

• The COVID-19 pandemic necessitates rapid development of vaccines and therapeutics.
• The SARS-CoV-2 spike protein is a critical target but is metastable and challenging for recombinant production.
• Existing methods face limitations in producing stable, high-yield spike proteins.

Purpose of the Study:

• To engineer a stabilized prefusion spike protein of SARS-CoV-2 with improved yield and stability.
• To identify beneficial mutations for enhancing spike protein expression and robustness.
• To facilitate the development of effective vaccines and diagnostics.

Main Methods:

• Structure-guided design of 100 spike protein variants.
• Identification and combination of 26 beneficial individual substitutions.
• Characterization of the HexaPro variant using cryo-electron microscopy.
• Assessment of HexaPro's stability under various stress conditions.

Main Results:

• Identified 26 substitutions that enhance spike protein yield and stability.
• Developed HexaPro, a variant with six proline substitutions, achieving 10-fold higher expression.
• HexaPro demonstrated stability against heat, room temperature storage, and freeze-thaw cycles.
• Cryo-EM confirmed HexaPro retains the prefusion spike conformation at 3.2 angstrom resolution.

Conclusions:

• High-yield production of a stabilized prefusion spike protein is achievable.
• HexaPro represents a significant advancement for SARS-CoV-2 vaccine and diagnostic development.
• This stabilized protein accelerates research and development for combating COVID-19.