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Specific interaction between anticodon nuclease and the tRNA(Lys) wobble base
Y Jiang1, R Meidler, M Amitsur
1Department of Biochemistry, Tel Aviv University, Ramat Aviv 69978, Israel.
Abstract:
The bacterial tRNA(Lys)-specific PrrC-anticodon nuclease cleaves its natural substrate 5' to the wobble base, yielding 2',3'-cyclic phosphate termini. Previous work has implicated the anticodon of tRNA(Lys) as a specificity element and a cluster of amino acid residues at the carboxy-proximal half of PrrC in its recognition. We further examined these assumptions by assaying unmodified and hypomodified derivatives of tRNA(Lys) as substrates of wild-type and mutant alleles of PrrC. The data show, first, that the anticodon sequence and wobble base modifications of tRNA(Lys) play major roles in the interaction with anticodon nuclease. Secondly, a specific contact between the substrate recognition site of PrrC and the tRNA(Lys) wobble base is revealed by PrrC missense mutations that suppress the inhibitory effects of wobble base modification mutations. Thirdly, the data distinguish between the anticodon recognition mechanisms of PrrC and lysyl-tRNA synthetase.
Insights
The bacterial PrrC nuclease requires specific tRNA(Lys) anticodon sequences and wobble base modifications for cleavage. Mutations reveal direct interactions between PrrC and the tRNA wobble base.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- The PrrC enzyme is a bacterial tRNA(Lys)-specific anticodon nuclease.
- It cleaves its substrate 5' to the wobble base, producing cyclic phosphate termini.
- Previous studies suggested tRNA(Lys) anticodon and PrrC's carboxy-proximal residues are key for recognition.
Purpose of the Study:
- To investigate the roles of tRNA(Lys) anticodon sequence and wobble base modifications in PrrC recognition.
- To identify specific interactions between PrrC and the tRNA(Lys) wobble base.
- To differentiate PrrC's anticodon recognition from that of lysyl-tRNA synthetase.
Main Methods:
- Assaying unmodified and hypomodified tRNA(Lys) derivatives with wild-type and mutant PrrC alleles.
- Analyzing PrrC missense mutations that suppress wobble base modification effects.
Main Results:
- The anticodon sequence and wobble base modifications of tRNA(Lys) significantly influence PrrC interaction.
- PrrC missense mutations indicate a direct contact between the enzyme's recognition site and the tRNA(Lys) wobble base.
- Distinct anticodon recognition mechanisms were observed for PrrC and lysyl-tRNA synthetase.
Conclusions:
- Both tRNA(Lys) anticodon sequence and wobble base modifications are critical for PrrC binding and cleavage.
- A direct physical interaction occurs between the PrrC enzyme and the tRNA(Lys) wobble base.
- PrrC exhibits a unique mechanism for anticodon recognition compared to other tRNA-binding proteins like lysyl-tRNA synthetase.