Improving the accuracy of macromolecular structure refinement at 7 Å resolution

Area of Science:

• Structural Biology
• X-ray Crystallography

Background:

• Molecular replacement and refinement in X-ray crystallography are difficult at low resolution.
• Initial models can deviate significantly from the true high-resolution structure.

Purpose of the Study:

• To compare different refinement methods for low-resolution X-ray crystallography data.
• To evaluate the effectiveness of Direct Electron Nucleus (DEN) refinement against standard methods.

Main Methods:

• Utilized synchrotron diffraction data of photosystem I at 7.4 Å resolution.
• Compared standard refinement with DEN refinement using various initial models of increasing structural deviation.
• Assessed model quality using R(free) values, atomic root-mean-square differences, and electron density connectivity.

Main Results:

• Standard refinement worsened initial models and increased R(free) values.
• DEN refinement improved even the most deviated starting models, as indicated by lower R(free) and better structural accuracy.
• The optimal protocol involved DEN refinement following initial segmented rigid-body refinement.
• The fraction of atoms within 2 Å of the true structure improved from 24% to 60% for the most distant initial model.

Conclusions:

• DEN refinement is a superior method for improving low-resolution crystallographic models compared to standard refinement.
• R(free) values remain a useful indicator of model accuracy even at low resolutions.
• The findings suggest DEN refinement protocols can significantly enhance structural determination from low-resolution data.