High-Resolution Electron Diffraction of Hydrated Protein Crystals at Room Temperature

Area of Science:

• Structural biology
• Biophysics
• Materials science

Background:

• Protein structural characterization is vital for understanding function.
• Electron crystallography offers advantages over X-ray diffraction for small crystals.
• Existing electron crystallography methods are limited by crystal fragility and cryogenic requirements.

Purpose of the Study:

• To develop a room-temperature methodology for preserving protein nanocrystals for electron diffraction.
• To overcome the limitations of crystal fragility and cryogenic techniques in electron crystallography.
• To enable in situ structural studies of hydrated nanocrystals.

Main Methods:

• Hermetically sealing protein crystals in their native solution using graphene-coated grids.
• Employing a low-dose data collection strategy to minimize electron radiation damage.
• Utilizing a hybrid-pixel direct electron detector for data acquisition.

Main Results:

• Successfully preserved lysozyme nanocrystals in their natural environment at room temperature.
• Obtained diffraction patterns with reflections up to 3 Å resolution.
• Successfully indexed diffraction data using a template-matching algorithm, demonstrating feasibility of in situ electron diffraction.

Conclusions:

• The developed methodology enables in situ protein electron diffraction at room temperature.
• This approach offers a viable alternative to cryogenic techniques for structural biology.
• The method is applicable to a wide range of hydrated nanocrystals for various research and technological applications.