Multi-particle cryo-EM refinement with M visualizes ribosome-antibiotic complex at 3.5 Å in cells

Area of Science:

• Structural Biology
• Biophysics
• Computational Biology

Background:

• High-resolution macromolecular structure determination using cryo-electron microscopy (cryo-EM) requires precise alignment and correction of sample dynamics.
• In-cell structural biology presents unique challenges due to sample motion and three-dimensional deformations during data acquisition.

Purpose of the Study:

• To introduce M, a novel software tool for cryo-electron microscopy data processing.
• To develop a unified framework for refining cryo-EM data, incorporating spatial deformation models and aberration correction.
• To enable high-resolution structural determination of macromolecules within intact cells.

Main Methods:

• Development of M, a reference-based, multi-particle refinement framework for cryo-EM data.
• Integration of a comprehensive spatial deformation model and in silico correction of electron-optical aberrations within M.
• Application of M in conjunction with Warp and RELION for processing cryo-EM data from intact bacterial cells.

Main Results:

• M provides a unified optimization framework for both frame-series and tomographic tilt-series data.
• Demonstrated that tilt-series data can achieve resolutions comparable to frame-series data for purified protein specimens.
• Achieved residue-level resolution of a 70S ribosome bound to an antibiotic within intact bacterial cells.

Conclusions:

• M significantly advances cryo-EM data processing by addressing spatial deformations and aberrations.
• The computational tool facilitates high-resolution structural biology studies in cellular environments.
• This work overcomes previous resolution limitations in tomographic cryo-EM data analysis.