Table 1.
Establishing the rotational status of mutant ribosomes
| Region | rRNA base | Non-rotated | A106R | S104D | Rotated |
|---|---|---|---|---|---|
| B7a | A2207 | 2 | 2 | 4 | 4 |
| G913 (SSU) | 1 | 2 | 4 | 4 | |
| B2a | U2258 | 2 | 2 | 3 | 4 |
| A2262 | 0 | 0 | 1 | 2 | |
| C1644 (SSU) | 1 | 1 | 2 | 2 | |
| G1645 (SSU) | 0 | 0 | 2 | 1 | |
| B3 | U2301 | 0 | 0 | 1 | 1 |
| G2302 | 0 | 0 | 0 | 0 | |
| A1655 (SSU) | 1 | 2 | 2 | 2 | |
| U1656 (SSU) | 2 | 2 | 2 | 2 | |
| AC | U2860 | 1 | 2 | 0 | 0 |
| U2924 | 4 | 2 | 0 | 1 | |
| A2926 | 4 | 2 | 1 | 1 | |
| SRL | U3023 | 1 | 1 | 0 | 0 |
| A3027 | 2 | 3 | 0 | 0 |
Non-rotated control yeast ribosomes were primed with polyU and contained Ac-Phe-tRNAPhe in the P-site. Rotated control ribosomes were primed with polyU, loaded with deacylated Phe-tRNA and incubated with eEF-2-GDPNP. These complexes, along with salt-washed vacant Rpl10-A106R and Rpl10-S104D ribosomes were chemically probed with 1M7 and analyzed by hSHAPE. B7, B2a and B3 denote the corresponding intersubunit bridges. Numerical values reflect statistically normalized reactivities on a scale of 0–4, with 4 being most reactive (18,19).