Switching recognition of two tRNA synthetases with an amino acid swap in a designed peptide.
The genetic code is based on specific interactions between transfer RNA (tRNA) synthetases and their cognate tRNAs. The anticodons for methionine and isoleucine tRNAs differ by a single nucleotide, and changing this nucleotide in an isoleucine tRNA is sufficient to change aminoacylation specificity to methionine. Results of combinatorial mutagenesis of an anticodon-binding-helix loop peptide were used to design a hybrid sequence composed of amino acid residues from methionyl- and isoleucyl-tRNA synthetases. When the hybrid sequence was transplanted into isoleucyl-tRNA synthetase, active enzyme was generated in vivo and in vitro. The transplanted peptide did not confer function to methionyl-tRNA synthetase, but the substitution of a single amino acid within the transplanted peptide conferred methionylation and prevented isoleucylation. Thus, the swap of a single amino acid in the transplanted peptide switches specificity between anticodons that differ by one nucleotide.
Auld
D S
DS
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139, USA.
Schimmel
P
P
D000595:Amino Acid Sequence; D000926:Anticodon; D005816:Genetic Complementation Test; D018257:Helix-Loop-Helix Motifs; D007533:Isoleucine-tRNA Ligase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008718:Methionine-tRNA Ligase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism; D012355:RNA, Transfer, Ile / Q000378:metabolism*; D012358:RNA, Transfer, Met / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D046249:Transfer RNA Aminoacylation
D000926:Anticodon; D009842:Oligopeptides; D012355:RNA, Transfer, Ile; D012358:RNA, Transfer, Met; D011994:Recombinant Proteins; D008718:Methionine-tRNA Ligase; D007533:Isoleucine-tRNA Ligase