Chapter 11Synthesis and Processing of the Proteome

Aminoacyl-tRNA synthetases attach amino acids to tRNAs

The attachment of amino acids to tRNAs - ‘charging’ in molecular biology jargon - is the function of the group of enzymes called aminoacyl-tRNA synthetases. The chemical reaction that results in aminoacylation occurs in two steps. An activated amino acid intermediate is first formed by reaction between the amino acid and ATP, and then the amino acid is transferred to the 3′ end of the tRNA, the link being formed between the -COOH group of the amino acid and the -OH group attached to either the 2′ or 3′ carbon on the sugar of the last nucleotide, which is always an A (Figure 11.4).

Figure 11.4 Aminoacylation of a tRNA. The result of aminoacylation by a Class II aminoacyl-tRNA synthetase is shown, the amino acid being attached via its -COOH group to the 3′-OH of the terminal nucleotide of the tRNA. A Class I aminoacyl-tRNA synthetase attaches the amino acid to the 2′-OH group.

With a few exceptions, organisms have 20 aminoacyl-tRNA synthetases, one for each amino acid. This means that groups of isoaccepting tRNAs are aminoacylated by a single enzyme. Although the basic chemical reaction is the same for each amino acid, the 20 aminoacyl-tRNA synthetases fall into two distinct groups, Class I and Class II, with several important differences between them. In particular, Class I enzymes attach the amino acid to the 2′-OH group of the terminal nucleotide of the tRNA, whereas Class II enzymes attach the amino acid to the 3′-OH group (Ibba et al., 2000).

Aminoacylation must be carried out accurately: the correct amino acid must be attached to the correct tRNA if the rules of the genetic code are to be followed during protein synthesis. It appears that an aminoacyl-tRNA synthetase has high fidelity for its tRNA, the result of an extensive interaction between the two, covering some 25 nm² of surface area and involving the acceptor arm and anticodon loop of the tRNA, as well as individual nucleotides in the D and TΨC arms. The interaction between enzyme and amino acid is, of necessity, less extensive, amino acids being much smaller than tRNAs, and presents greater problems with regard to specificity because several pairs of amino acids are structurally similar. Errors do therefore occur, at a very low rate for most amino acids but possibly as frequently as one aminoacylation in 80 for difficult pairs such as isoleucine and valine. Most errors are corrected by the aminoacyl-tRNA synthetase itself, by an editing process that is distinct from aminoacylation, involving different contacts with the tRNA (Hale et al., 1997; Silvian et al., 1999).

In most organisms, aminoacylation is carried out by the process just described, but a few unusual events have been documented. These include a number of instances where the aminoacyl-tRNA synthetase attaches the incorrect amino acid to atRNA, this amino acid subsequently being transformed into the correct one by a second, separate chemical reaction. This was first discovered in the bacterium Bacillus megaterium for synthesis of glutamine-tRNA^Gln (i.e. glutamine attached to its tRNA). This aminoacylation is carried out by the enzyme responsible for synthesis of glutamic acid-tRNA^Glu, and initially results in attachment of a glutamic acid to the tRNA^Gln (Figure 11.5A). This glutamic acid is then converted to glutamine by transamidation catalyzed by a second enzyme. The same process is used by various other bacteria (although not Escherichia coli) and by the archaea. Some archaea also use transamidation to synthesize asparagine-tRNA^Asn from aspartic acid-tRNA^Asn (Ibba et al., 2000). In both of these cases, the amino acid that is synthesized by the modification process is one of the 20 that are specified by the genetic code. There are also two examples where the modification results in an unusual amino acid. The first example is the conversion of methionine to N-formylmethionine (Figure 11.5B), producing the special aminoacyl-tRNA used in initiation of bacterial translation. The second example occurs in both prokaryotes and eukaryotes and results in synthesis of selenocysteine, which is specified in a context-dependent manner by some 5′-UGA-3′ codons. These codons are recognized by a special tRNA^SeCys, but there is no aminoacyl-tRNA synthetase that is able to attach selenocysteine to this tRNA. Instead, the tRNA is aminoacylated with a serine by the seryl-tRNA synthetase, and then modified by replacement of the -OH group of the serine with an -SeH, to give selenocysteine (Figure 11.5C; Low and Berry, 1996).

Figure 11.5.  Unusual types of aminoacylation. (A) In some bacteria, tRNA^Gln is aminoacylated with glutamic acid, which is then converted to glutamine by transamidation. (B) The special tRNA used inin initiation of translation in bacteria is aminoacylated with methionine, which is then converted to N-formylmethionine. (C) tRNASeCys in various organisms is initially aminoacylated with serine.