Dynamics of EF-G interaction with the ribosome explored by classification of a heterogeneous cryo-EM dataset.

A method of supervised classification using two available structure templates was applied to investigate the possible heterogeneity existing in a large cryo-EM dataset of an Escherichia coli 70S ribosome-EF-G complex. Two subpopulations showing the ribosome in distinct conformational states, related by a ratchet-like rotation of the 30S subunit with respect to the 50S subunit, were extracted from the original dataset. The possible presence of additional intermediate states is discussed.

[1]  M. S. Chapman,et al.  Study of the Structural Dynamics of the E. coli 70S Ribosome Using Real-Space Refinement , 2003, Cell.

[2]  J. Frank,et al.  The ribosome at improved resolution: new techniques for merging and orientation refinement in 3D cryo-electron microscopy of biological particles. , 1994, Ultramicroscopy.

[3]  J. Frank.  Ribosomal dynamics explored by cryo-electron microscopy. , 2001, Methods.

[4]  R. Craig,et al.  Capturing time-resolved changes in molecular structure by negative staining. , 2003, Journal of Structural Biology.

[5]  J. Frank,et al.  Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. , 2001, Molecular cell.

[6]  Joachim Frank,et al.  Locking and Unlocking of Ribosomal Motions , 2003, Cell.

[7]  J. Frank,et al.  A cryo-electron microscopic study of ribosome-bound termination factor RF2 , 2003, Nature.

[8]  Måns Ehrenberg,et al.  Peptidyl-tRNA Regulates the GTPase Activity of Translation Factors , 2003, Cell.

[9]  E. Egelman,et al.  Flexibility of the rings: structural asymmetry in the DnaB hexameric helicase. , 2002, Journal of molecular biology.

[10]  M. Ehrenberg,et al.  Novel roles for classical factors at the interface between translation termination and initiation. , 1999, Molecular cell.

[11]  Joachim Frank,et al.  Cryo‐EM reveals an active role for aminoacyl‐tRNA in the accommodation process , 2002, The EMBO journal.

[12]  Alasdair C. Steven,et al.  Dynamics of herpes simplex virus capsid maturation visualized by time-lapse cryo-electron microscopy , 2003, Nature Structural Biology.

[13]  J. Trinick,et al.  A computer-controlled spraying-freezing apparatus for millisecond time-resolution electron cryomicroscopy. , 1998, Journal of structural biology.

[14]  Joachim Frank,et al.  A ratchet-like inter-subunit reorganization of the ribosome during translocation , 2000, Nature.

[15]  George Harauz,et al.  Self-organisation of cryoelectron micrographs of the phosphoenolpyruvate synthase from Staphylothermus marinus , 1998 .

[16]  Scott M Stagg,et al.  Incorporation of aminoacyl-tRNA into the ribosome as seen by cryo-electron microscopy , 2003, Nature Structural Biology.

[17]  A Leith,et al.  SPIDER and WEB: processing and visualization of images in 3D electron microscopy and related fields. , 1996, Journal of structural biology.