Sub-2 Angstrom resolution structure determination using single-particle cryo-EM at 200 keV.

Area of Science:

• Structural Biology
• Biophysics
• Microscopy

Background:

• Direct electron detectors (DEDs) and software advancements have improved single-particle cryogenic electron microscopy (cryo-EM) resolution.
• A gap exists between current cryo-EM resolutions and the theoretical limits of transmission electron microscopes (TEMs).
• 200 kV TEMs are typically used for sample screening, not high-resolution structure determination.

Purpose of the Study:

• To demonstrate that 200 kV TEMs can achieve high-resolution cryo-EM structure determination.
• To highlight the underutilization of 200 kV TEMs for advanced structural biology applications.
• To showcase the capability of a specific 200 kV TEM (Talos Arctica) for high-resolution cryo-EM.

Main Methods:

• Utilized a 200 kV Talos Arctica transmission electron microscope equipped with a K2 direct electron detector.
• Applied single-particle cryo-electron microscopy techniques to biological specimens.
• Processed and refined cryo-EM data to achieve high-resolution reconstructions.

Main Results:

• Achieved a resolution of approximately 1.7 Å for macromolecular structures.
• Identified ordered water molecules and distinguished holes in aromatic residues at this resolution.
• Demonstrated the capability of the 200 kV TEM for high-resolution cryo-EM.

Conclusions:

• 200 kV electrons are suitable for high-resolution single-particle cryo-EM.
• 200 kV TEMs can be effectively utilized for determining macromolecular structures.
• This approach has implications for structure-based drug design and other applications.