The Escherichia coli large ribosomal subunit at 7.5 A resolution.

Background: In recent years, the three-dimensional structure of the ribosome has been visualised in different functional states by single-particle cryo-electron microscopy (cryo-EM) at 13-25 A resolution. Even more recently, X-ray crystallography has achieved resolution levels better than 10 A for the ribosomal structures of thermophilic and halophilic organisms. We present here the 7.5 A solution structure of the 50S large subunit of the Escherichia coli ribosome, as determined by cryo-EM and angular reconstitution.

Results: The reconstruction reveals a host of new details including the long alpha helix connecting the N- and C-terminal domains of the L9 protein, which is found wrapped like a collar around the base of the L1 stalk. A second L7/L12 dimer is now visible below the classical L7/L12 'stalk', thus revealing the position of the entire L8 complex. Extensive conformational changes occur in the 50S subunit upon 30S binding; for example, the L9 protein moves by some 50 A. Various rRNA stem-loops are found to be involved in subunit binding: helix h38, located in the A-site finger; h69, on the rim of the peptidyl transferase centre cleft; and h34, in the principal interface protrusion.

Conclusions: Single-particle cryo-EM is rapidly evolving towards the resolution levels required for the direct atomic interpretation of the structure of the ribosome. Structural details such as the minor and major grooves in rRNA double helices and alpha helices of the ribosomal proteins can already be visualised directly in cryo-EM reconstructions of ribosomes frozen in different functional states.