Maximizing the potential of electron cryomicroscopy data collected using direct detectors

Area of Science:

• Structural Biology
• Biophysics
• Electron Microscopy

Background:

• Single-particle electron cryomicroscopy (cryo-EM) is advancing rapidly due to direct electron detector technology.
• Direct detectors offer superior detective quantum efficiency and readout speeds compared to older technologies like film and CCDs.
• This advancement facilitates higher resolution imaging of biological macromolecules.

Purpose of the Study:

• To evaluate the potential of the Gatan K2 Summit direct detector for near-atomic resolution cryo-EM.
• To assess this potential when coupled with a mid-range transmission electron microscope (FEI TF20 Twin).
• To compare data quality with a high-end microscope setup (FEI Titan Krios with Falcon I detector).

Main Methods:

• Utilized a Gatan K2 Summit direct electron detector on an FEI TF20 Twin transmission electron microscope.
• Employed motion correction and dose fractionation strategies to mitigate beam-induced motion and stage drift.
• Reconstructed a 3D map of Sulfolobus turreted icosahedral virus (STIV) as a test specimen.

Main Results:

• Achieved a 4.4Å resolution 3D map of STIV, demonstrating the capability of the system.
• Investigated various motion correction and dose fractionation techniques, detailing their impact on resolution.
• Compared the obtained data quality against that from a high-end FEI Titan Krios microscope.

Conclusions:

• The Gatan K2 Summit direct detector, when used with a mid-range TEM, can achieve high-resolution cryo-EM reconstructions.
• Careful selection of motion correction and dose fractionation protocols is crucial for optimal resolution.
• This study provides valuable insights into the performance of direct detectors on accessible instrumentation.