Codon discrimination and anticodon structural context.

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Abstract
Site-directed mutagenesis has been used to change the nucleotide C in the wobble position of tRNA1Gly (CCC) to U. The mutated tRNA was tested for its ability to read glycine codons in an in vitro protein-synthesizing system programmed with the phage message MS2-RNA that had been modified by site-directed mutagenesis so as to make it possible to monitor conveniently the reading of all four glycine codons. The results showed that while the efficiency of tRNA1Gly (UCC) was comparable to that of mycoplasma tRNAGly (UCC) in the reading of the codon GGA, the mycoplasma tRNAGly was far more efficient than the tRNA1Gly (UCC) in the reading of the codons GGU and GGC. Thus, the anticodon UCC, when present in the structural context of the tRNA1Gly molecule, behaved as predicted by the wobble rules while in the structural context of the mycoplasma tRNAGly it read without discrimination between the nucleotides in the third codon position, in violation of the wobble restrictions. The result with the codon GGG showed that the anticodon UCC, when present in tRNA1Gly, was considerably less efficient in reading this codon than it was in the structural context of the mycoplasma tRNAGly. It would therefore seem that the anticodon UCC, when present in a certain tRNA, can be an efficient wobbler, while in the molecular environment of another tRNA it is markedly restricted in its ability to wobble.