Making routine native SAD a reality: lessons from beamline X06DA at the Swiss Light Source

Area of Science:

• Macromolecular crystallography
• Structural biology
• Biophysics

Background:

• Native single-wavelength anomalous dispersion (SAD) phasing is a preferred de novo method utilizing intrinsic anomalous scattering.
• Successful native SAD experiments rely on precise reflection intensity measurements and careful data collection.
• Low Bijvoet ratios and complex crystal symmetries can present challenges in experimental phasing.

Purpose of the Study:

• To review and report on the low-dose, multiple-orientation native SAD phasing protocol at beamline X06DA (PXIII).
• To demonstrate the method's utility and success rate in solving de novo macromolecular structures.
• To highlight the application of native SAD phasing in challenging crystallographic cases.

Main Methods:

• Implementation of a low-dose, multiple-orientation data collection strategy for native SAD phasing.
• Application of the protocol to conventional crystals (>50 µm) over a four-year period.
• Case study involving native SAD phasing on a streptavidin-biotin crystal with P21 symmetry and a low Bijvoet ratio (0.6%).

Main Results:

• The native SAD phasing protocol has been successfully applied to solve 45 de novo structures, with 13 published.
• The method has become the primary experimental phasing choice for users at beamline X06DA.
• Demonstrated success in phasing a challenging crystal with low Bijvoet ratio and P21 symmetry.

Conclusions:

• The reviewed low-dose, multiple-orientation native SAD protocol is experimentally simple, fast, and effective.
• Native SAD phasing is a reliable and increasingly popular method for de novo structure determination in macromolecular crystallography.
• The method's versatility extends to challenging cases, and intrinsic anomalous signals can aid model building and ion assignment.