RT Journal Article SR Electronic T1 Multi-particle cryo-EM refinement with M visualizes ribosome-antibiotic complex at 3.7 Å inside cells JF bioRxiv FD Cold Spring Harbor Laboratory SP 2020.06.05.136341 DO 10.1101/2020.06.05.136341 A1 Tegunov, Dimitry A1 Xue, Liang A1 Dienemann, Christian A1 Cramer, Patrick A1 Mahamid, Julia YR 2020 UL http://biorxiv.org/content/early/2020/06/05/2020.06.05.136341.abstract AB Cryo-electron microscopy (cryo-EM) enables macromolecular structure determination in vitro and in situ. In addition to aligning individual particles, accurate registration of sample motion and 3D deformation during exposures is crucial for achieving high resolution. Here we describe M, a software tool that establishes a reference-based, multi-particle refinement framework for cryo-EM data and improves the results of structure determination. M provides a unified optimization framework for both in vitro frame series and in situ tomographic tilt series data. We show that tilt series data can provide the same resolution as frame series, indicating that the alignment step no longer limits the resolution obtainable from tomographic data. In combination with Warp and RELION, M improves upon previous methods, and resolves a 70S ribosome bound to an antibiotic inside bacterial cells at a nominal resolution of 3.7 Å. Thus, computational tools are now available to resolve structures from tomographic in situ cryo-EM data at residue level.Competing Interest StatementThe authors have declared no competing interest.