DECODING WITH THE A-I WOBBLE PAIR IS INEFFICIENT

tRNAs with inosine (I) in the first position read three codons ending in U, C and A, However, A-ending codons read with I are rarely used, In Escherichia coli, CGA/U/C are all read solely by tRNA(lCG)(Arg). CGU and CGC are very common codons, but CGA is very rare, Three independent in vivo assays show that translation of CGA is relatively inefficient, In the first, nine tandem CGA cause a strong rho-mediated polar effect on expression of a lacZ reporter gene, The inhibition is made more extreme by a mutation in ribosomal protein S12 (rpsL), which indicates that ribosomal binding by tRNA(ICG)(Arg) is slow and/or unstable in the CGA cluster, The second assay, in which codons are substituted for the regulatory UGA of the RF2 frameshift, confirms that aa-tRNA selection is slow and/or unstable at CGA. In the third assay, CGA is found to be a poor 5' context for amber suppression, which suggests that an A:I base pair in the P site can interfere with translation of a codon in the A site. Two possible errors, frameshifting and premature termination by RF2, are not significant causes for inefficiency at CGA. It is concluded that the A:I pair destabilizes codon:anticodon complexes during two successive ribosomal cycles, and it is suggested that these properties contribute to the rare usage of codons read with the A:I base pair.