Quantitative comparison of zero-loss and conventional electron diffraction from two-dimensional and thin...

Area of Science:

• Structural biology
• Electron crystallography
• Biophysics

Background:

• Electron crystallography offers higher resolution for biological macromolecules than X-ray crystallography due to larger scattering cross-sections.
• Inelastic electron scattering poses a significant challenge, degrading data quality in electron crystallography.
• Smaller and thinner protein crystals are viable for electron crystallography, enabling analysis of challenging samples.

Purpose of the Study:

• To evaluate the impact of energy filtering on electron diffraction data quality.
• To assess the effectiveness of removing inelastically scattered electrons for structural analysis.
• To determine the utility of energy filtering for both 2-D and 3-D crystal datasets.

Main Methods:

• Collected electron diffraction data from thin 3-D and 2-D protein crystals at 93 K.
• Utilized an energy filtering electron microscope operating at 200 kV accelerating voltage.
• Analyzed data with and without energy filtering to assess the removal of inelastically scattered electrons.

Main Results:

• Removal of inelastically scattered electrons significantly improved intensity data statistics and R(Friedel) factor up to 3-Å resolution.
• Energy filtering demonstrated a prominent effect on thicker crystals and was also significant for thin crystals.
• Filtered data sets exhibited superior intensity statistics compared to unfiltered data collected at higher voltage (300 kV) and lower temperature (4 K).

Conclusions:

• Energy filtering is an effective tool for enhancing electron crystallography data quality.
• This technique is particularly valuable for the structure analysis of thin 3-D and 2-D crystals.
• Energy filtering improves data statistics, especially when data are collected at high tilt angles.