The importance of alignment accuracy for molecular replacement.

Area of Science:

• Structural biology
• Computational biology
• Biophysics

Background:

• Molecular replacement (MR) is a key technique for determining protein structures.
• Automated software for MR exists, but optimal search model preparation is underexplored.
• Advances in sequence comparison enable better detection and alignment of distant homologs.

Purpose of the Study:

• To investigate if simple homology models from improved sequence alignments enhance MR.
• To assess the impact of different alignment methods on MR success rates.

Main Methods:

• A parallelized pipeline was used for MR, including homology detection and model preparation.
• Search models were generated using standard (BLAST) and advanced (PSI-BLAST, FFAS) alignment methods.
• 27 crystal structures were determined and analyzed.

Main Results:

• Models based on more accurate alignments showed higher success rates.
• This improvement was particularly notable for structures with <35% sequence identity to the search model.
• The MR pipeline facilitated all steps from detection to rebuilding.

Conclusions:

• Simple homology models, when based on accurate sequence alignments, significantly increase MR success.
• Utilizing advanced alignment techniques is beneficial for MR, especially for distantly related homologs.