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Evidence for a unique first position codon-anticodon mismatch in vivo
M J Toth1, E J Murgola, P Schimmel
1Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Journal of Molecular Biology
|May 20, 1988
Summary
Researchers investigated beta-lactamase gene mutations, finding that a specific glycine substitution (SG68(GGC)) unexpectedly produced low activity. This activity resulted from rare mistranslation of the genetic code, a novel observation in molecular biology.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- The Ser68(AGC) codon in the beta-lactamase gene was mutated to glycine codons (GGA and GGC).
- Glycine at position 68 normally results in inactive beta-lactamase due to the absence of a nucleophilic side-chain essential for the reaction mechanism.
Purpose of the Study:
- To investigate the phenotypic differences between two 'same-sense' beta-lactamase mutants, SG68(GGA) and SG68(GGC).
- To determine the cause of residual beta-lactamase activity in the SG68(GGC) mutant by examining the role of translational fidelity.
Main Methods:
- Mutant beta-lactamase alleles (SG68(GGA) and SG68(GGC)) were introduced into Escherichia coli strains with altered translational fidelity (rpsD and rpsL mutations).
- Beta-lactamase activity and protein production were assessed in these modified strains using a maxi-cell system.
Main Results:
- The SG68(GGA) allele showed no detectable beta-lactamase activity, while the SG68(GGC) allele produced minimal activity.
- A mutation decreasing translational fidelity (rpsD) significantly increased activity for the SG68(GGC) allele, but not the SG68(GGA) allele.
- A mutation increasing translational fidelity (rpsL) had minimal effect on the SG68(GGC) allele's activity.
Conclusions:
- The SG68(GGC) allele's activity is attributed to a rare mistranslation event (approximately 0.1%) by tRNA(GCU)Ser, leading to the production of active beta-lactamase.
- This study provides the first evidence of an A/G wobble at the first position of the codon-anticodon interaction involving a wild-type tRNA.
- Differential effects of translational fidelity mutations on the SG68(GGC) allele highlight the sensitivity of genetic code decoding.