Modulation of Decoding Fidelity by Ribosomal Proteins S4 and S5

Ribosomal proteins S4 and S5 participate in the decoding and assembly processes on the ribosome and the interaction with spe-cific antibiotic inhibitors of translation. Many of the characterized mutations affecting these proteins decrease the accuracy of translation, leading to a ribosomal-ambiguity phenotype. Structural analyses of ribosomal complexes indicate that the tRNA selection pathway involves a transition between the closed and open conformations of the 30S ribosomal subunit and requires disruption of the interface between the S4 and S5 proteins. In agreement with this observation, several of the mutations that pro-mote miscoding alter residues located at the S4-S5 interface. Here, the Escherichia coli rpsD and rpsE genes encoding the S4 and S5 proteins were targeted for mutagenesis and screened for accuracy-altering mutations. While a majority of the 38 mutant pro-teins recovered decrease the accuracy of translation, error-restrictive mutations were also recovered; only a minority of the mu-tant proteins affected rRNA processing, ribosome assembly, or interactions with antibiotics. Several of the mutations affect resi-dues at the S4-S5 interface. These include five nonsense mutations that generate C-terminal truncations of S4. These truncations are predicted to destabilize the S4-S5 interface and, consistent with the domain closure model, all have ribosomal-ambiguity phenotypes. A substantial number of the mutations alter distant locations and conceivably affect tRNA selection through indi-rect effects on the S4-S5 interface or by altering interactions with adjacent ribosomal proteins and 16S rRNA. Cellular protein synthesis systems translatemRNAs quickly andwith high accuracy. The accuracy of the decoding process can, however, be altered by agents such as the antibiotic streptomyci