SEQUENCE SLIDER: expanding polyalanine fragments for phasing with multiple side-chain hypotheses

Area of Science:

• Structural biology
• Computational biology
• Biophysics

Background:

• Fragment-based molecular replacement (MR) is a powerful technique for solving macromolecular structures.
• Challenges arise at low resolution, with low solvent content, high beta-sheet composition, or low fragment accuracy.
• Existing methods like ARCIMBOLDO struggle in these difficult scenarios.

Purpose of the Study:

• To introduce SEQUENCE SLIDER, a novel method to overcome limitations in fragment-based MR.
• To extend initial polyalanine fragments with side chains in a multisolution framework.
• To improve the accuracy and success rate of macromolecular structure determination.

Main Methods:

• SEQUENCE SLIDER extends initial fragments with side chains using a multisolution approach.
• Fragment selection is guided by log-likelihood gain (LLG) calculated with Phaser.
• Sequence assignment is based on secondary structure prediction or alignment with homologs.
• Brute-force trial of sequence hypotheses through side-chain building and refinement.

Main Results:

• SEQUENCE SLIDER successfully addressed challenging cases, including low-resolution data and low solvent content.
• The method was validated on test cases and previously unknown structures.
• Demonstrated success in solving the structures of MltC and a 638-residue pneumococcal lipoprotein.

Conclusions:

• SEQUENCE SLIDER significantly enhances fragment-based MR, particularly for difficult datasets.
• The method provides a robust framework for side-chain extension and sequence assignment.
• Enables the determination of complex macromolecular structures previously intractable with standard methods.