Self-assembled superstructure alleviates air-water interface effect in cryo-EM

Area of Science:

• Structural Biology
• Biophysics
• Materials Science

Background:

• Cryo-electron microscopy (cryo-EM) is crucial for atomic-resolution protein structure determination.
• Protein adsorption to the air-water interface during specimen preparation hinders cryo-EM success and resolution.
• Preferential orientation of proteins is a significant challenge in cryo-EM data analysis.

Purpose of the Study:

• To develop a robust method for high-resolution cryo-EM analysis by addressing air-water interface issues.
• To investigate the efficacy of impenetrable surfactant monolayers in passivating air-water interface problems.
• To enable the structural determination of proteins, including small ones, at near-atomic resolution.

Main Methods:

• Development and application of graphene-surfactant self-assembled monolayers (GSAMs).
• Utilizing GSAMs to control protein interaction with the air-water interface during cryo-EM specimen preparation.
• Employing GSAMs to enrich particle orientations and improve particle utilization ratios.

Main Results:

• Facilitated a 3.3-Å resolution reconstruction of the 100-kDa ACE2-RBD protein complex, overcoming preferential orientation.
• Enabled successful near-atomic resolution structure determination for small proteins (<100 kDa).
• Demonstrated improved particle orientation and utilization for multiple protein types.

Conclusions:

• GSAMs provide an effective solution to air-water interface challenges in cryo-EM.
• This method significantly enhances the success rate and achievable resolution in cryo-EM studies.
• The findings offer a key strategy for controlling protein-interface interactions in structural biology.