Annealing macromolecular crystals.

Area of Science:

• Crystallography
• Structural Biology
• Materials Science

Background:

• Macromolecular crystals often exhibit poor diffraction quality after flash cooling due to high mosaicity or ice formation.
• Crystals with unsatisfactory diffraction are typically discarded, leading to data loss.

Purpose of the Study:

• To present crystal annealing as a method to improve diffraction quality in flash-cooled macromolecular crystals.
• To describe protocols for annealing crystals to enhance their suitability for diffraction experiments.

Main Methods:

• Flash-cooled crystals with poor diffraction are removed from the goniometer.
• Crystals are incubated in cryoprotectant buffer at room or growth temperature for approximately 3 minutes.
• Crystals are then remounted and flash-cooled again. In situ annealing involves warming the crystal on the loop before re-cooling.

Main Results:

• Crystal annealing successfully improves diffraction quality from crystals that would otherwise be discarded.
• The technique is effective in reducing mosaicity and mitigating diffraction from ice.
• In situ annealing shows promise for crystals with low solvent content or small solvent channels.

Conclusions:

• Crystal annealing is a valuable post-flash-cooling treatment for improving macromolecular crystal diffraction.
• This method enhances the usability of crystals for structural determination.
• Variations in annealing protocols, including in situ methods, offer flexibility for different crystal properties.