Post-transcriptional Modifications of the tRNA Anticodon Stem and Loop (ASL) Affect the Ability of tRNA to Bind Synonymous Codons.

Abstract

The Genetic Code is arranged into sixteen codon boxes, where the four codons in each box are similar in their first two letters but differ at the third position (the wobble position). In the universal Genetic Code, each amino acid, except for Tryptophan and Methionine that have one codon each, is encoded by two to six codons (two to six-fold degenerate). There are fewer tRNA species than codons; therefore, a tRNA species can recognize more than one codon. This flexibility in recognition resides in the third position of the codon:anticodon pairing, the wobble position. Codon recognition by tRNA is dependent on the anticodon loop. The sequence of the anticodon (tRNA positions 34, 35, and 36) does not necessarily predict codon binding according to Watson-Crick rules. In all organisms, post-transcriptional modifications occur quite extensively and of great variety at the anticodon loop. These modifications, usually found on the nucleosides in tRNA position 34 (the wobble position) and position 37, direct the tRNA’s ability to read codons accurately and efficiently. Just as the types of modifications are diverse, the abilities of modifications to recognize codons also vary. When a particular amino acid is encoded by an entire codon box, such as four-fold degenerate Valine, we show that a tRNA species with a specific modification can read all four codons of that box. The modification 5-oxyacetic acid (cmo5) at the wobble position of tRNAVal (tRNAVal-cmo5U34) allows cmo5U34 to recognize U, A, G, and perhaps C. In instances where the difference between the codes of two amino acids resides only in the third letter of their codons (2-fold degenerate codons in a split box), modifications at the wobble position of the anticodon restrict codon recognition to the two codons specific for that 2-fold degenerate amino acid. For example, Lysine has two codons (AAA and AAG) that share a codon box with Asparagine codons (AAU and AAC). The modified nucleoside 5-methoxycarbonylmethyl-2-thiouridine at the wobble position of human tRNA (tRNALys-mcm5s2U34) confers this tRNA’s ability to restrict codon recognition to the two Lysine codons only. Similar to the cytoplasmic tRNAs, mitochondrial tRNAs also contain posttranscriptional modifications. The mitochondria deviate from the universal Genetic Code in that it uses the universal Isoleucine codon AUA to decode Methionine. In all organisms, there are two Methionine tRNAs: an initiator tRNAMet and an elongator tRNAMet. Mitochondria, however, have but one tRNAMet that acts as both initiator and elongator, has characteristics of both types of tRNAMet, and decodes AUG and AUA in the aminoacyl- (entry or A)-site and the peptidyl (P)-site of the ribosome. The human mitochondrial tRNAMet is modified with a 5-formyl-group at the wobble position cytidine-34 (hmtRNAMetf5C34). This modification allows the hmtRNAMet f5C34 to expand codon recognition to include AUA in translating Methionine. At times, the mitochondria use Isoleucine codons AUU and AUC to initiate translation. Surprisingly, the 5-formyl modification of hmtRNAMet-f5C34 also allows codon-reading expansion at the P-site to include the entire codon box AUN.