The termination of translation occurs when a ribosome that is translating an mRNA encounters a stop codon. Since there are no tRNAs that can base pair with stop codons, the A-site of a ribosome that is positioned at a stop codon is able to bind a protein called release factor, which fosters hydrolysis of the bond that links the growing polypeptide to the tRNA occupying the ribosome’s P-site (see 8.27 in Slonczweski text).
But what if there was a tRNA that could base pair with a stop codon? Binding of this tRNA at the A-site would block release factor binding and, if the tRNA was “charged” with an amino acid, elongation of the growing polypeptide could continue. It turns out that particular point mutations in the anticodon loops of certain tRNAs can interfere with translational termination in just this way. Mutant tRNAs of this kind are encountered in nature and are referred to as suppressor tRNAs (for their ability to suppress translational termination).
A single base substitution in the anticodon loop of a tRNA for glutamic acid (tRNAglu) might allow this mutant tRNA to base pair with certain termination codons and suppress translational termination and also cause a glutamic acid residue into the growing polypeptide at certain stop codons.
Assume that a certain strain of E. coli contains two copies of a gene for tRNAglu and that one of these tRNA genes has a point mutation in the region encoding its anticodon loop that converts its gene product into a suppressor tRNA. Assume also that the wildtype tRNAglu has a U residue at the wobble position and, due to the relaxed base pairing rules that occur only at the wobble position, this single tRNA can base pair with both Glu codons.
A) What is the nucleotide sequence of the wildtype anticodon of tRNAglu ?
(write the sequence in 5’è3′ polarity)
Wildtype anticodon sequence:
B) A single point mutation in this anticodon can convert it to one with suppressor activity. What is the mutant anticodon sequence?
(write the sequence in 5’è3′ polarity)
Mutant anticodon sequence:
C) At which stop codons would the mutant tRNAglu be able to suppress translational termination? (remember that G-U base pairing is possible at the wobble position).
D) Why would this point mutation be lethal if there were only a single gene for tRNAglu?
E) Mutations of the sort described above are rarely encountered in nature, due in part to the existence of a key step in the chain of events that helps ensure translational fidelity. What is this step and how might it prevent suppressor tRNAs from being able to incorporate amino acids at stop codons?.
F) Are there any other tRNAs for other amino acids that might be converted into suppressor tRNAs by point mutations in their anticodon loops? Give one example.
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