Initiator tRNA molecules

Initiator tRNA Molecules and Selection of Initiation Codon... 

The initiator tRNA molecule in prokaryotes—tRNA — has several properties that distinguish it from all other tRNA molecules. One feature is that the tRNA is first acylated with methionine, and then the methionine is modified. Acylation is by methionyl tRNA synthetase, which also charges tRNA . However, the methionine of charged tRNA is immediately recognized by another enzyme, tRNA methionyl transformylase, which transfers a formyl group from N °-formyltetrahydrofolate (fTHF) to the amino group of the methionine to form... 

Eukaryotic initiator tRNA molecules differ from the prokaryotic initiator molecule in several ways. The most striking difference is that whereas eukaryotic organisms produce both a normal tRNA and an initiator tRNA, which is also charged with methionine, the methionine does not undergo formylation. In eukaryotes, the first amino acid in a growing polypeptide chain is Met and not fMet. The codon for both kinds of tRNA molecules in eukaryotes is AUG, just as for prokaryotes. 

The translation of the mRNA into proteins is the final step in the biological flow of information (see Fig. 6.1). Similar to other macromolecular polymerizations, protein synthesis can be divided into initiation, chain elongation, and termination. Critical players in this process are the aminoacyl transfer RNAs (tRNAs). These molecules form the interface between the mRNA and the growing polypeptide. Activation of tRNA involves the addition of an amino acid to its acceptor stem, a reaction catalyzed by an aminoacyl-tRNA synthetase. Each aminoacyl-tRNA synthetase is highly specific for one amino acid and its corresponding tRNA molecule. The anticodon loop of each aminoacyl-tRNA interacts... 

At one point or another during protein synthesis, several other proteins will be associated with the ribosome. These include factors that help in initiating the synthetic process, others that help in elongating the peptide chain, and yet others that play a role in terminating the synthesis of a peptide chain. Beyond this, there is also the mRNA to consider, as well as the aminoacylated tRNA molecules. Finally, since protein biosynthesis consumes energy, there is the hydrolysis of ATP and GTP to AMP and GDP, respectively, by the ribosome. 

In all tRNAs the bases can be paired to form "clover-leaf" structures with three hairpin loops and sometimes a fourth as is indicated in Fig. 5-30.329 331 This structure can be folded into the L-shape shown in Fig. 5-31. The structure of a phenylalanine-carrying tRNA of yeast, the first tRNA whose structure was determined to atomic resolution by X-ray diffraction, is shown.170/332 334 An aspartic acid-specific tRNA from yeast,335 and an E. coli chain-initiating tRNA, which places N-formyl-methionine into the N-terminal position of proteins,336,337 have similar structures. These molecules are irregular bodies as complex in conformation as globular proteins. Numerous NMR studies show that the basic... 

Discrimination between some pairs of tRNAs depends entirely on the anticodon sequence. For example, tRNAMet contains the anticodon CAU. That for a minor tRNAIle is the same except that the cytosine has been posttranscriptionally modified by covalent linkage of a molecule of lysine via its e-amino group to C2 of the cytosine. The latter base (Iysidine) is correctly recognized by E. coli isoleucyl-tRNA synthetase but, if the cytosine is unmodified, it is aminoacylated by methionyl-tRNA synthetase.192 In most instances the acceptor specificity, or tRNA identity, is not determined solely by the anticodon sequence. Thus, when a methionine initiator tRNA was modified to contain a tryptophan anticodon, it was only partially charged with tryptophan in vivo. However, when A73 of the methionine tRNA was also converted to G73, only tryptophan was inserted.193 Nucleotide 73 (Fig. 

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. 

Formylation of methionylated tRNA " allows differentiation of the AUG start codon from internal AUG codons (14). MetRS aminoacylates tRNA" with methionine. A formyl group is linked covalently to the charged methionine via its amino moiety by the methionyl-tRNA formyltransferase enzyme, which uses N -formyl tetrahydrofolate as the formyl donor. This fMet-tRNA molecule binds directly to the P site of the ribosome to initiate protein synthesis, as compared with the A-site to which elongator tRNAs bind. 

When the 70S initiation complex has been formed, the ribosome is ready for the elongation phase of protein synthesis. The fMet-tRNAf molecule occupies the P site on the ribosome. The other two sites for tRNA molecules, the A site and the E site, are empty. Formylmethionyl-tRNAf is positioned so that its anticodon pairs with the initiating AUG (or GUG) codon on mRNA. This interaction sets the reading frame for the translation of the entire mRNA. 

These products can be formed by attack of OH on quinonoid intermediates of the type postulated for thymidylate S5mthetase (formed in step d of Fig. 15-21). A related reaction is the posttranscriptional conversion of a single uracil residue in tRNA molecules of some bacteria to a thymine ring (a ribothymidylic acid residue). In this case, FADH2 is used as a reducing agent to convert the initial adduct of Fig. 15-21 to the ribothymidylic acid and THF. "... 

Figure 12.4 outlines the process of protein synthesis involving the ribosome, mRNA, a series of amino-acyl transfer RNA (tRNA) molecules (at least one for each amino acid) and accessory protein factors involved in initiation, elongation and termination. As the process is essentially the same in prokaryotic (bacterial) and eukaryotic cells (i.e. higher organisms and mammalian cells) it is surprising that there are so many selective agents which act in this area (see Fig. 12.1). 

The amino group of the aminoacyl-tRNA in the A site is well positioned to attack the ester linkage between the initiator tRNA and the formylmethionine molecule (Figure 30.20). T he peptidyl transferase center includes bases that promote this reaction by helping to form an —NTI2 group on the A-site aminoacyl-tRNA and by helping to stabilize the tetrahedral intermediate... 

---