tRNA hopping: effects of mutant tRNAs.
ABSTRACT Movement of tRNA and mRNA through the ribosome is coupled. However, selection for suppression of a -1 frameshift mutation in Escherichia coli has yielded a class of mutant tRNAs that can violate this mechanism and "hop" or disengage from their cognate codons and re-pair downstream in the mRNA. Previously described tRNA mutants of this class included those with insertions in the anticodon of tRNA(Val)1. This report describes further tRNA(Val)1 alterations that enhance hopping; these include a novel insertion in the anticodon loop, base substitutions in the anticodon stem and a base deletion in the variable loop. These results indicate that several different features of a tRNA are important for maintaining stable codon-anticodon interactions and coupled movement of tRNA and mRNA during the elongation phase of protein synthesis.
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ABSTRACT: Mutants of translation components which compensate for both -1 and +1 frameshift mutations showed the first evidence for framing malleability. Those compensatory mutants isolated in bacteria and yeast with altered tRNA or protein factors are reviewed here and are considered to primarily cause altered P-site realignment and not altered translocation. Though the first sequenced tRNA mutant which suppressed a +1 frameshift mutation had an extra base in its anticodon loop and led to a textbook "yardstick" model in which the number of anticodon bases determines codon size, this model has long been discounted, although not by all. Accordingly, the reviewed data suggest that reading frame maintenance and translocation are two distinct features of the ribosome. None of the -1 tRNA suppressors have anticodon loops with fewer than the standard seven nucleotides. Many of the tRNA mutants potentially affect tRNA bending and/or stability and can be used for functional assays, and one has the conserved C74 of the 3' CCA substituted. The effect of tRNA modification deficiencies on framing has been particularly informative. The properties of some mutants suggest the use of alternative tRNA anticodon loop stack conformations by individual tRNAs in one translation cycle. The mutant proteins range from defective release factors with delayed decoding of A-site stop codons facilitating P-site frameshifting to altered EF-Tu/EF1alpha to mutant ribosomal large- and small-subunit proteins L9 and S9. Their study is revealing how mRNA slippage is restrained except where it is programmed to occur and be utilized.Microbiology and molecular biology reviews: MMBR 04/2009; 73(1):178-210. · 12.59 Impact Factor
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ABSTRACT: If a ribosome shifts to an alternative reading frame during translation, the information in the message is usually lost. We have selected mutants of Salmonella typhimurium with alterations in tRNA(cmo5UGG)(Pro) that cause increased frameshifting when present in the ribosomal P-site. In 108 such mutants, two parts of the tRNA molecule are altered: the anticodon stem and the D-arm, including its tertiary interactions with the variable arm. Some of these alterations in tRNA(cmo5UGG)(Pro) are in close proximity to ribosomal components in the P-site. The crystal structure of the 30S subunit suggests that the C-terminal end of ribosomal protein S9 contacts nucleotides 32-34 of peptidyl-tRNA. We have isolated mutants with defects in the C-terminus of S9 that induce +1 frameshifting. Combinations of changes in tRNA(cmo5UGG)(Pro) and S9 suggest that an interaction occurs between position 32 of the peptidyl-tRNA and the C-terminal end of S9. Together, our results suggest that the cause of frameshifting is an aberrant interaction between the peptidyl-tRNA and the P-site environment. We suggest that the "ribosomal grip" of the peptidyl-tRNA is pivotal for maintaining the reading frame.Journal of Molecular Biology 12/2008; 385(2):350-67. · 3.91 Impact Factor
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ABSTRACT: The hypothesis that tRNA sidearm loops bear anticodons assumes crossovers between anticodon and sidearms, or translation by expressed aminoacylated tRNA halves forming single stem-loops. Only the latter might require ribosomal adaptations. Drosophila mitochondrial codon usages coevolve with sidearm numbers bearing matching putative anticodons (comparing different codon families in one genome, macroevolution) and when comparing different genomes for single codon families (microevolution). Coevolution between Drosophila and yeast mitochondrial antisense tRNAs and codon usages partly confounds microevolutionary patterns for putative sidearm anticodons. Some tRNA sidearm loops have more than seven nucleotides, putative expanded anticodons potentially matching quadruplet codons (tetracodons, codons expanded by a fourth silent position, forming tetragenes (predicted by alignment analyses of Drosophila mitochondrial genomes)). Tetracodon numbers coevolve with expanded tRNA sidearm loops. Sidearm coevolution with amino acid usages and tetragenes occurs for putative anticodons in 5' and 3' sidearms loops (D and TΨC loops, respectively), are stronger for the D-loop. Results slightly favour isolated stem-loops upon crossover hypotheses. An alternative hypothesis, that patterns observed for sidearm 'anticodons' do not imply translational activity, but recognition signals for tRNA synthetases that aminoacylate tRNAs, is incompatible with tetracodon/tetra-anticodon coevolution. Hence analyses strengthen translational hypotheses for tRNA sidearm anticodons, tetragenes, and antisense tRNAs.Journal of Theoretical Biology 09/2013; · 2.35 Impact Factor