Volume 1

Gene expression / Benjamin Lewin.

  • Lewin, Benjamin.
Date:
1974-1977
Catalogue details

Licence: Attribution-NonCommercial 4.0 International (CC BY-NC 4.0)

Credit: Gene expression / Benjamin Lewin. Source: Wellcome Collection.

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    in figure 1.13, This protein has B activity, so that its B segment is presumably unaffected by the attachment of the A region. The nature of mutants in the rll cistron can therefore be tested by placing them in series with r 1589 by genetic recombination. Testing the double mutant for B protein activity shows whether the mutant in A is of the missense or nonsense type; a nonsense mutant allows no B activity, because synthesis of the polypeptide chain is terminated in the A region, but a missense mutation in the A region does not influence B activity. A protein B protein Tinnnry mm Single protein with B activity TÏ5WWW Protein with B activity despite^ mutation in A segmenta- Normal phage r1589 mutant : separation region deleted No protein corresponding to B segment nrcnr Missense mutation Nonsense mutation Figure 1.13: The rl589 test for nonsense mutations. In normal phage, the A and B cistrons each produce a separate protein. The rl589 deletion synthesises a single protein representing both cistrons but which has the activity characteristic of the B protein. A missense mutation in the A region does not affect the activity of the B region of the combined protein. But the presence of a nonsense mutation terminates protein synthesis in the A region so that protein for the B region is no longer made The behaviour of the phage nonsense mutants isolated by this test depends upon which strain of E.coli is used as the bacterial host; in non-permissive strains the phage cannot grow because the protein chain is prematurely terminated, but growth can occur in other, permissive, (or suppressor ) strains. Benzer and Champe suggested that this dependence on the host bacterial strain can be accounted for if the suppressor strain contains an unusual (mutant) aminoacyl-tRNA which responds to the nonsense codon by inserting an amino acid into the growing polypeptide chain, whereas the non-permissive strain lacks this species and terminates protein synthesis at the nonsense codon. Nucleotide Sequences of Nonsense Codons Nonsense mutants can be classified according the suppressor loci to which they respond. They fall into three classes. The mutants found by Benzer and Champe all respond to the same suppressor and have been termed amber. Brenner and Beckwith (1965) found another class, termed ochre , which are not suppressed by amber suppressors. However, suppressors of the ochre mutants also suppress amber mutants; this implies that, although the two