Translation codon-anticodon interactions

The 3 terminal redundancy of the genetic code and its mechanistic basis were first appreciated by Francis Crick in 1966. He proposed that codons and anticodons interact in an antiparallel manner on the ribosome in such a way as to require strict Watson-Crick pairing (that is, A-U and G-C) in the first two positions of the codon but to allow other pairings in its 3 terminal position. Nonstandard base pairing between the 3 terminal position of the codon and the 5 terminal position of the anticodon alters the geometry between the paired bases Crick s proposal, labeled the wobble hypothesis, is now viewed as correctly describing the codon-anticodon interactions that underlie the translation of the genetic code. 

The number of mammalian mitochondrial tRNA molecules is 22, which is less than the minimum number (32) needed to translate the universal code. This is possible because in each of the fourfold redundant sets—e.g., the four alanine codons GCU, GCC, GCA, and GCG—only one tRNA molecule (rather than two, as explained above) is used. In each set of four tRNA molecules, the base in the wobble position of the anticodon is U or a modified U (not I). It is not yet known whether this U is base-paired in the codon-anticodon interaction or manages to pair weakly with each of the four possible bases. For those codon sets that are doubly redundant—e.g., the two histidine codons CAU and CAC—the wobble base always forms, a G-U pair, as in the universal code. The structure of the human mitrochondrial tRNA molecule is also different from that of the standard tRNA molecule (except for mitochondrial tRNA UUX). (X = any nucleotide.) The most notable differences are the following ... 

The first two base pairings in a codon-anticodon interaction confer most of the specificity required during translation. Recall that most redundant codons specifying a certain amino acid possess identical nucleotides in the first two positions. These interactions are standard (i.e., Watson-Crick) base pairings. 

Initiation. Translation begins with initiation, when the small ribosomal subunit binds an mRNA. The anticodon of a specific tRNA, referred to as an initiator tRNA, then base pairs with the initiation codon AUG. Initiation ends as the large ribosomal subunit combines with the small subunit. There are two sites on the complete ribosome for codon-anticodon interactions the P (peptidyl) site (now occupied by the enitiator tRNA) and the A (aminoacyl) site. In both prokaryotes and eukaryotes, mRNAs are read simultaneously by numerous ribosomes. An mRNA with several ribosomes bound to it is referred to as a polysome. In actively growing prokaryotes, for example, the ribosomes attached to an mRNA molecule may be separated from each other by as few as 80 nucleotides. 

Selection of the specific aminoacyl-tRNA to be bound at the ribosomal A site is by base-pairing between the relevant mRNA codon and the tRNA anticodon. Because this interaction involves only a triplet of bases and hence a maximum of nine hydrogen bonds (see Fig. IB), it is intrinsically unstable at physiological temperatures and is probably stabilized by components of the ribosome to allow sufficient time for peptide bond synthesis to occur. Also, the codon-anticodon pairing must be monitored for fidelity in order to minimize errors in translation. In E. coli there is genetic and biochemical evidence that one of the proteins of the small ribosomal subunit, S12, is involved in ensuring the fidelity of normal translation and in causing the mistranslation which occurs in the presence of the antibiotic streptomycin due to incorrect codon-anticodon interactions. 

After translocation the ribosomal P site is occupied by dipeptidyl-tRNA and the vacant A site contains the third mRNA codon. Entry of the next aminoacyl-tRNA, selected as before by the codon-anticodon interaction, into the A site (Fig. If) enables peptide bond synthesis to continue and repeated operation of the elongation-translocation cycle gives rise to a stepwise elongation of the nascent polypeptide chain, each complete cycle elongating the chain by one amino acid residue and moving the mRNA by one codon in the 5 to 3 direction. When the end of the coding sequence is reached and one of the termination (or stop) codons has entered the A site, translation stops and the completed polypeptide chain is released. 

According to their theory, as the polycistronic messenger RNA moves in relation to the polysome system, the velocity of protein synthesis in its various parts is slowed. They postulated that the sequence of the genes in the histidine operon (which does not correspond to the biochemical sequence of reactions) is connected with the number of molecules of each enzyme synthesized. By analyzing the frequency of mutations of polarity, they concluded that many triplets (of the 64 possible) can retard the transcription and translation of information. The essence of the matter is that if any nucleotide triplet (codon) XYZ requires an anticodon in the molecules of acceptor sRNA for itstranslationinto a protein "text," a lowered content of this fraction of sRNA with the corresponding anticodon may act as modulator of the velocity of translation, which is reduced at this locus in connection with a decrease in the number of codon-anticodon interactions. 

Initiation Factors 2 and 3 have seemingly opposing functions. While IF2 promotes the binding of tRNA to the 308 subunit, IF3 can be considered the subunit antiassociation factor because it increases the rates of subunit exchange and complex dissociation. In fact the two functions cooperate to facilitate formation of the correct 308 initiation complex— IF2 preferentially enhances the binding of the amino-blocked initiator tRNA and IF3 specifically increases the rate of noninitiator tRNA dissociation from the ternary complex. Initiation Factor 3 also contributes to the fidelity of translation by confirming the codon-anticodon interaction on the 308 subunit. 

Figure 10 Alteration of the genetic code for incorporation of non-natural amino acids, (a) In nonsense suppression, the stop codon UAG is decoded by a non-natural tRNA with the anticodon CUA. In vivo decoding of the UAG codon by this tRNA is in competition with termination of protein synthesis by release factor 1 (RFl). Purified in vitro translation systems allow omission of RF1 from the reaction mixture, (b) A new codon-anticodon pair can be created using four-base codons such as GGGU. Crystal structures of these codon-anticodon complexes in the ribosomal decoding center revealed that the C in the third anticodon position interacts with both the third and fourth codon position (purple line) while the extra A in the anticodon loop does not contact the codon.(c) Non-natural base pairs also allow creation of new codon-anticodon pairs. Shown here is the interaction of the base Y with either base X or (hydrogen bonds are indicated by red dashes).

Tricyclic hypermodified nucleosides are found in archaeal and eukaryotic tRNAs and are frequently observed at position 34 (wobble base) or position 37 (adjacent to the anticodon). Position 37 typically contains a hypermodified nucleoside such as N -threonylcarbamoyladenosine (t A), 2-methylthio-N -isopentenyl-ade-nosine (ms i A-37), or wybutosine (yW). yW and its derivatives occur at position 37 in archaeal and eukaryotic phenylalanine tRNA (tRNAphe). The modifications serve to maintain the correct translational reading frame via hydrophobic interactions, which reinforce codon—anticodon pairing and prevent incorrect Watson—Crick base-pairing. Studies have shown that unmodified tRNA leads to translational defects that have been implicated in different pathological states. ... 

Several fluorescence and biochemical experiments reveal the detailed mechanism of translation and the contribution of KPR towards the fidelity of protein synthesis. The incorporation of an amino acid into the peptide is composed of two consecutive processes initial selection of tRNA at the A site of the ribosome followed by KPR [23]. Various factors affect the initial selection of tRNA such as the HB energy between the codon-anticodon base pairs, the specific interactions between the large subunit of the ribosome and aa-tRNA, etc. The contribution of the initial selection step to the overall error fraction for Escherichia coli is observed to be -1/6, compared to the overall error fraction -7 x 10 for cognate and near-cognate anticodons. As a consequence the contribution of KPR is expected to be 1/24 i.e. -80% of the observed fidelity comes from the KPR. The translation process occurs through the following mechanism (see Figure 13.2). 

In the translation process, codons in mRNA and anticodons in tRNA interact in an antiparallel manner such that the two 5 bases of the codon interact with two... 

At the ribosome, which travels along the mRNA, the tRNA molecule is bound such that its anticodon can interact with a nucleotide triplet on mRNA (the codon). If the anticodon is complementary to a codon triplet on the mRNA, the amino acid attached at the 3 -terminus of the tRNA is transferred to the amino terminus of the growing polypeptide chain if it is not complementanty, the tRNA is rejected and another one is checked for complementary. The whole process is repeated until the synthesis of the protein is completed. It is initiated, as well as terminated, by specific codons regulating this translation. 

---