Table 2.
UGA readthrough activity of E. coli SelB variants containing amino-acid exchanges in the aminoacyl-tRNA-binding pocketa
| Readthrough activity (Miller units) with |
|||
|---|---|---|---|
| Plasmid | Mutations | Ser-tRNASec | Sec-tRNASec |
| pMB | Wild type | 8.9 | 273.4 |
| pMT7 | Empty vector | 1.5 | 0.6 |
| pMBD-1 | Y42H | 7.9 | 397.7 |
| pMBD-2 | D180E | 4.4 | 375.7 |
| pMBD-3 | R181D | 0.8 | 232.0 |
| pMBD-4 | R236N | 6.2 | 334.0 |
| pMBD-12 | Y42H, D180E | 3.1 | 469.4 |
| pMBD-13 | Y42H, R181D | 1.0 | 336.4 |
| pMBD-14 | Y42H, R236N | 4.2 | 163.8 |
| pMBD-23 | D180E, R181D | 0.5 | 15.2 |
| pMBD-24 | D180E, R236N | 6.0 | 495.1 |
| pMBD-34 | R181D, R236N | 0.5 | 0.7 |
| pMBD-123 | Y42H, D180E, R181D | 0.9 | 214.9 |
| pMBD-124 | Y42H, D180E, R236N | 21.8 | 544.2 |
| pMBD-134 | Y42H, R181D, R236N | 0.4 | 0.0 |
| pMBD-234 | D180E, R181D, R236N | 0.7 | 0.2 |
| pMBD-1234 | Y42H, D180E, R181D, R236N | 0.9 | 1.4 |
| aControl plasmids or plasmids bearing mutated selB alleles were transferred into AF90422/pWT and PT91300/pWT to measure UGA readthrough with Ser-tRNASec and Sec-tRNASec, respectively, and β-galactosidase activities were determined. The results of at least three independent experiments, each of which was performed in triplicate, were averaged. | |||