Aminoacyl-tRNA synthetase, use

Some aminoacyl-tRNA synthetases use the anticodon of the tRNA as a recognition site as they attach the amino acid to the hydroxyl group at the 3 -end of the tRNA (Fig. 15.6). However, other synthetases do not use the anticodon but recognize only bases located at other positions in the tRNA. Nevertheless, insertion of the amino acid into a growing polypeptide chain depends solely on the bases of the anticodon, through complementary base-pairing with the mRNA codon. 

Their DNA is circular. Their ribosomes are more like those of bacteria than those of eukaryotes. Their aminoacyl-tRNA synthetases use bacterial tRNAs but not eukaryotic tRNAs. In general, they do not have introns in their genomes. Their mRNA uses a Shine-Dalgarno sequence. 

Gunasekera, N., Lee, S. W., Kim, S., Musier-Forsyth, K., and Arriaga, E., Nuclear localization of aminoacyl-tRNA synthetases using single-ceU capillary electrophoresis laser-induced fluorescence analysis. Ana/. Chem., 76, 4741 746, 2004. 

Adenosine 5">triphosphate (ATP) regeneration systems acetate kinase, use, 112,114,115/ aminoacyl-tRNA synthetase, use, 116.118f/,119... 

Cech s group was the first to have success in this direction (Piccirilli, 1992). Using a genetically modified Tetrahymena ribozyme, they were able to hydrolyse an ester bond between the amino acid A-formylmethionine and the corresponding tRNAf Met. The reaction was, however, very slow, only about 5 to 15 times faster than the uncatalysed reaction. The authors ventured to suggest that these ribozymes could have functioned as the first aminoacyl tRNA synthetases. 

The formation of 0-seryl or 0-prolyl esters (Figure 1) of certain N-hydroxy arylamines has been inferred from the observations that highly reactive intermediates can be generated in vitro by incubation with ATP, serine or proline, and the corresponding aminoacyl tRNA synthetases (11,12,119). For example, activation of N-hydroxy-4-aminoquinoline-l-oxide (119,120), N-hydroxy-4-aminoazobenzene (11) and N-hydroxy-Trp-P-2 (121) to nucleic acid-bound products was demonstrated using seryl-tRNA synthetase from yeast or rat ascites hepatoma cells. More recently, hepatic cytosolic prolyl-, but not seryl-, tRNA synthetase was shown to activate N-hydroxy-Trp-P-2 (12) however, no activation was detectable for the N-hydroxy metabolites of AF, 3,2 -dimethyl-4-aminobiphenyl, or N -acetylbenzidine (122). 

Although aminoacyl-tRNA synthetases are necessary for protein synthesis in all tissues, their importance in chemical carcinogenesis is difficult to assess. Mutation induction by this pathway has been studied extensively (123), yet metabolic activation in a carcinogen-target tissue has not been demonstrated. The only exception is hepatic prolyl-tRNA synthetase activation of N-hydroxy-Trp-P-2 however, hepatic O-acetylation of this substrate also occurs to an appreciable extent (12). Further investigations involving the use of specific enzyme inhibitors would be helpful in addressing this problem. 

Figure 27-16 summarizes what we know about the nucleotides involved in recognition by some aminoacyl-tRNA synthetases. Some nucleotides are conserved in all tRNAs and therefore cannot be used for discrimination. 

FIGURE 27-16 Nucleotide positions in tRNAs that are recognized by aminoacyl-tRNA synthetases. Some positions (blue dots) are the same in all tRNAs and therefore cannot be used to discriminate one from another. Other positions are known recognition points for one (orange) or more (green) aminoacyl-tRNA synthetases. Structural features other than sequence are important for recognition by some of the synthetases. 

Importance of the Second Genetic Code Some aminoacyl-tRNA synthetases do not recognize and bind the anticodon of their cognate tRNAs but instead use other structural features of the tRNAs to impart binding specificity. The tRNAs for alanine apparently fall into this category. 

The aminoacyl-tRNA synthetases join amino acids to their appropriate tRNA molecules for protein synthesis. They have the very important task of selecting both a specific amino acid and a specific tRNA and joining them. The enzymes differ in size and other properties. However, they all appear to function by a common basic chemistry that makes use of cleavage of ATP at Pa (Eq. 12-48) via an intermediate aminoacyl adenylate and that is outlined also in Eq. 17-36. These enzymes are discussed in Chapter 29. ... 

While peptide antibiotics are synthesized according to enzyme-controlled polymerization patterns, both proteins and nucleic acids are made by template mechanisms. Tire sequence of their monomer emits is determined by genetically encoded information. A key reaction in the formation of proteins is the transfer of activated aminoacyl groups to molecules of tRNA (Eq. 17-36). Tire tRNAs act as carriers or adapters as explained in detail in Chapter 29. Each aminoacyl-tRNA synthetase must recognize the correct tRNA and attach the correct amino acid to it. The tRNA then carries the activated amino acid to a ribosome, where it is placed, at the correct moment, in the active site. Peptidyltransferase, using a transacylation reaction, in an insertion mechanism transfers the C terminus of the growing peptide chain onto the amino group of... 

One advantage of this method is that the enzyme is in contact with any particular substrate molecule for a short time only, and so can be used in cases in which the enzyme slowly hydrolyzes or chemically transforms the ligand. Another advantage is that some available gels are able to distinguish between the size of one polymer and another, so that, for example, the binding of a tRNA (Mr = 25 000) to an aminoacyl-tRNA synthetase (Mr = 100 000) may be measured. 

Tsao ML, Tian F, Schultz P. Incorporation of unnatural reactive amino acids into phage display libraries using ortiiogonal mutant aminoacyl-tRNA synthetases for post-translational modification. PCT Int. Appl. 2007 114. 

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