Ribosomal decoding processes at codons in the A or P sites depend differently on 2'-OH groups

The importance of 2'-OH groups of codons for binding of cognate tRNAs to ribosomal P and A sites was analyzed applying the following strategy. An mRNA of 41 nucleotides was synthesized with the structure C16-GAA-UUC-GUC-C16 coding for glutamic acid (E), phenylalanine (F) and valine (V), respectively, in the middle (EFV-mRNA). A second template, the E(dF)V-mRNA, was identical except that it carried a deoxyribo-codon-dUdUdC- for phenylalanine. tRNA binding to the P site is totally insensitive to the presence or absence of the 2'-OH group of the P-site codon, and tRNA binding to the P site is also not affected if the A-site codon lacks the 2'-OH groups. However, binding is impaired if the deoxy-codon is present at the E site. In sharp contrast, the A-site binding of Ac-aminoacyl-tRNA was severely reduced in the presence of the deoxy-codon at the A site as well as at the P site. The results demonstrate that the correctness of base pairing is also "sensed" via a correct sugar structure of the codon, e.g. positioning of the sugar pucker (2'-OH), during the decoding process at the A site (elongation) but not during the decoding at the P site (initiation).