Extending the scope of coiled-coil crystal structure solution by AMPLE through improved ab initio modelling.

Area of Science:

• Structural Biology
• X-ray Crystallography
• Computational Biology

Background:

• The phase problem is a significant hurdle in solving protein structures via X-ray crystallography.
• Ab initio modeling and ideal secondary structures aid in solving novel protein structures without clear homologues.
• Highly helical structures, particularly coiled-coils, benefit from rigid alpha-helices as molecular replacement fragments.

Purpose of the Study:

• To expand the capabilities of the AMPLE program for solving coiled-coil protein structures.
• To improve ab initio modeling methods for enhanced structure solution.
• To enable coiled-coil structure determination at lower resolutions.

Main Methods:

• Implemented general pipeline improvements within the AMPLE program.
• Removed twin NCS (tNCS) correction in the molecular replacement process.
• Developed two enhanced ab initio modeling techniques: enforcing elongated helices and modeling oligomeric coiled-coils (dimers, trimers, tetramers).

Main Results:

• The new ab initio modeling approach overcame bias towards globular folds, enabling rapid, enhanced structure solution.
• Modeling of oligomeric coiled-coils and their use in molecular replacement improved success rates for difficult cases.
• Successfully determined parallel/antiparallel dimeric, trimeric, and tetrameric coiled-coil structures at resolutions as low as 3.3 Å.

Conclusions:

• The enhanced AMPLE program overcomes previous limitations, offering new functionality for coiled-coil structure solution.
• Automated modeling of elongated monomers and oligomers is now available in AMPLE via a 'coiled-coil' mode.
• These advancements facilitate protein structure determination for a wider range of coiled-coil proteins, especially at lower resolutions.