10.3.1.1—
tRNA
The code words (codons) of the genetic code were first established for E. coil. This work showed that there were 61 codons shared between twenty protein amino acids, i.e. there is more than one codon for some amino acids and these can attach to more than one tRNA species. Chemically different tRNA species which can be acylated by the same amino acid are called isoacceptor tRNAs.
However, it was soon realized that not every codon has a corresponding tRNA with a specific anticodon and that some tRNAs recognize more than one codon. Crick's 'wobble hypothesis' (1966) to account for this, proposed alternative base-pairing to occur between the base in the third position of the mRNA codon and the corresponding base in the tRNA anticodon. The proposed rules are set out in Fig. 10.5. Thus, inosine in the 5' position in the anticodon of an
Figure 10.5
The basis of the 'wobble' hypothesis. (X =
Any nucleotide; I = Inosine; G = Guanine;
U = Uridine; A = Adenine; C = Cytosine;)
(From Boulter et al. (Biol. Rev. 47, 113–75, 1972.))
aminoacyl-tRNA molecule allows this base to pair with either U, C or A in the third 3' position of a messenger codon. Similarly, with G or U in the 5' position in the anticodon, two codons may be recognized by a single aminoacyl-tRNA, whereas with C or A only a single codon is recognized. Since alternative base-pairing is only possible with the base in the third position of the codon, codons for the same amino acid which differ in either of the first two base positions, base-pair with different tRNAs. The only example of 'wobble' in the first base of a codon occurs with tRNAiMet , which recognizes both the initiator AUG or GUG codons (see later). Even so, separation of tRNAs by counter current-distribution, MAK columns, BD-cellulose columns and reversed phase chromatography has shown that there are more isoacceptor tRNAs than can be accounted for by the degeneracy of the genetic code. Similarly, isoacceptor tRNAs for amino acids exist in plants; for example, soyabean seedlings have at least six different tRNALeu species (Anderson & Cherry, 1969). Although few experiments have been carried out, it has been found in every case investigated that plants use the same codon in amino acid assignments as those of E. coli. However, plants may not use all of the possible codons for a particular amino acid, since some isoacceptor tRNA species may be absent, or because of the 'wobble' effect; e.g. Caskey et al., (1968) have shown that the isoleucyl tRNA from guinea pig liver recognizes AUU, AUC and AUA codons, whereas the corresponding isoleucyl-tRNA from E. coli only recognizes two of these, AUU and AUC. This response to different codons in the two organisms is explained if the liver tRNA had 5' inosine in the anticodon, while E. coli tRNA had guanine.
The whole question of the number and the activity of tRNAs in different tissues of plants is complicated by the fact that, (a) it is not certain whether the methods for separating tRNAs are completely effective; (b) it is known that during isolation and purification, partial modification of tRNA molecules can occur, which may affect their charging ability with amino acids or their ability to transfer amino acids to proteins, or both (see Sueoka & Kano-Sueoka, 1970); (c) the relative importance of the three protein synthesizing systems, cytoplasmic, chloroplast and mitochondrial, each with its associated tRNAs and synthetases may differ in different tissues. Nevertheless, different complements of isoacceptor tRNAs may occur in different tissues and changes in pattern have been demonstrated during development (Bick et al., 1970; Littauer & Inouye, 1973), and it has been suggested that isoacceptor tRNAs are involved in the control of protein synthesis. Garel et al., (1973) have shown that the complement of tRNAs synthesized in the silk gland is related to the mRNAs being translated and they have suggested defining this phenomenon as 'modulated tRNA biosynthesis'.