Selenium single-wavelength anomalous diffraction de novo phasing using an X-ray-free electron laser

Area of Science:

• Structural Biology
• Biophysics
• Biochemistry

Background:

• X-ray crystallography is the primary method for determining biological macromolecule structures.
• Challenges in X-ray crystallography include the phase problem and radiation damage.
• X-ray free electron lasers (XFELs) mitigate radiation damage but de novo phasing remains difficult.

Purpose of the Study:

• To demonstrate the feasibility of de novo phasing at X-ray free electron lasers (XFELs).
• To establish selenium anomalous diffraction (Se-SAD) as a viable phasing method at XFELs.
• To enable novel structure determination for macromolecules previously challenging to solve.

Main Methods:

• Utilized the Linac Coherent Light Source (LCLS), an X-ray free electron laser.
• Employed selenium anomalous diffraction (Se-SAD) at a single wavelength.
• Determined the structure of selenobiotinyl-streptavidin.

Main Results:

• Successfully obtained phases using Se-SAD at an X-ray FEL.
• Solved the structure of selenobiotinyl-streptavidin using XFEL-derived Se-SAD phases.
• Demonstrated that Se-SAD is a viable de novo phasing method at XFELs.

Conclusions:

• Selenium anomalous diffraction (Se-SAD) is now a practical phasing technique at X-ray free electron lasers (XFELs).
• This advancement overcomes previous limitations in de novo phasing at XFELs.
• Opens new avenues for atomic-resolution structure determination of biological macromolecules.