Fig. 5.
A previously unidentified translocation intermediate supports the asymmetric movement of the template–product duplex during translocation. (A) Stereo-pair images of the translocation intermediate complex C3S6 with a composite SA omit electron density map (contoured at 1.2 σ) of RNA, pyrophosphate (PPi), and residue D238 overlaid. The coloring scheme is as in Fig. 3A. The C3S4/5 pretranslocation (dark gray) and C3S1 posttranslocation (brown) complexes are shown with polymerases superimposed on the intermediate complex for comparisons. (B) RNA-only comparison of the translocation intermediate (Left: template in cyan; Right: product in green) and the pretranslocation complex (dark gray) in the same NAC. (C) Schematic illustration of the different protein–RNA interactions in the pretranslocation C3S4/5, intermediate C3S6, and posttranslocation C3S1 complexes. RNA movement of the C3S6 structure was estimated using the base, ribose, and phosphate components of the pre- and posttranslocation complexes as references. The zigzagged red symbol indicates the irregular backbone conformation of the template −2 position. Phosphate, ribose, and base are shown as circles, pentagons, and blocks, respectively. Solid arrows indicate hydrogen bonding, electrostatic, or hydrophobic interactions. Gray fonts indicate weaker interactions (judged by distance) compared with interactions involving the same residue in other structures. Underlining indicates a change of interaction partner(s) for a polymerase residue when switching from the posttranslocation complex to the intermediate complex. Strikethroughs indicate nonexistent interactions.