Single sequence of a helix-loop peptide confers functional anticodon recognition on two tRNA synthetases.
The specific aminoacylation of RNA oligonucleotides whose sequences are based on the acceptor stems of tRNAs can be viewed as an operational RNA code for amino acids that may be related to the development of the genetic code. Many synthetases also have direct interactions with tRNA anticodon triplets and, in some cases, these interactions are thought to be essential for aminoacylation specificity. In these instances, an unresolved question is whether interactions with parts of the tRNA outside of the anticodon are sufficient for decoding genetic information. Escherichia coli isoleucyl- and methionyl-tRNA synthetases are closely related enzymes that interact with their respective anticodons. We used binary combinatorial mutagenesis of a 10 amino acid anticodon binding peptide in these two enzymes to identify composite sequences that would confer function to both enzymes despite their recognizing different anticodons. A single peptide was found that confers function to both enzymes in vivo and in vitro. Thus, even in enzymes where anticodon interactions are normally important for distinguishing one tRNA from another, these interactions can be 'neutralized' without losing specificity of amino-acylation. We suggest that acceptor helix interactions may play a role in providing the needed specificity.
Auld
D S
DS
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Schmimmel
P
P
D000595:Amino Acid Sequence; D000926:Anticodon / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D014644:Genetic Variation; D018257:Helix-Loop-Helix Motifs / Q000235:genetics; D007533:Isoleucine-tRNA Ligase / Q000235:genetics* / Q000378:metabolism; D008718:Methionine-tRNA Ligase / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional
D000926:Anticodon; D004269:DNA, Bacterial; D008718:Methionine-tRNA Ligase; D007533:Isoleucine-tRNA Ligase