Double-flow focused liquid injector for efficient serial femtosecond crystallography

Area of Science:

• Biophysics
• Structural Biology
• Biochemistry

Background:

• Serial femtosecond crystallography (SFX) is crucial for determining protein structures.
• Efficient delivery of fresh protein crystals to the X-ray beam is a key challenge in SFX experiments.
• Existing nozzle technologies can be limited by sample consumption and jet stability.

Purpose of the Study:

• To develop and validate a novel double-flow focusing nozzle for SFX.
• To improve sample delivery efficiency and jet stability in X-ray free-electron laser (XFEL) experiments.
• To enable high-resolution structure determination of challenging protein targets at room temperature.

Main Methods:

• Introduction of a double-flow focusing nozzle design for SFX.
• Testing nozzle performance with various protein samples, including RNA polymerase II and extradiol ring-cleaving dioxygenase.
• Utilizing X-ray free-electron lasers (XFELs) for data collection.

Main Results:

• The double-flow focusing nozzle significantly reduces sample consumption compared to previous designs.
• Improved jet stability was observed, leading to more reliable crystal delivery.
• High-resolution, room-temperature structures of RNA polymerase II and an extradiol ring-cleaving dioxygenase were successfully determined.

Conclusions:

• The double-flow focusing nozzle is a significant advancement for SFX.
• This technology facilitates efficient structure determination of proteins at room temperature.
• The nozzle's improved performance opens new possibilities for studying protein dynamics and function.