Initiating translation with D-amino acids.
Here we report experimental evidence that the translation initiation apparatus accepts D-amino acids ((D)aa), as opposed to only L-methionine, as initiators. Nineteen (D)aa, as the stereoisomers to their natural L-amino acids, were charged onto initiator tRNA(fMet)(CAU) using flexizyme technology and tested for initiation in a reconstituted Escherichia coli translation system lacking methionine, i.e., the initiator was reprogrammed from methionine to (D)aa. Remarkably, all (D)aa could initiate translation while the efficiency of initiation depends upon the type of side chain. The peptide product initiated with (D)aa was generally in a nonformylated form, indicating that methionyl-tRNA formyltransferase poorly formylated the corresponding (D)aa-tRNA(fMet)(CAU). Although the inefficient formylation of (D)aa-tRNA(fMet)(CAU) resulted in modest expression of the corresponding peptide, preacetylation of (D)aa-tRNA(fMet)(CAU) dramatically increased expression level, implying that the formylation efficiency is one of the critical determinants of initiation efficiency with (D)aa. Our findings provide not only the experimental evidence that translation initiation tolerates (D)aa, but also a new means for the mRNA-directed synthesis of peptides capped with (D)aa or acyl-(D)aa at the N terminus.
Goto
Yuki
Y
Research Center of Advanced Science and Technology, The University of Tokyo, Tokyo, 153-8904, Japan.
Murakami
Hiroshi
H
Suga
Hiroaki
H
D000215:Acylation; D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D005821:Genetic Techniques; D007536:Isomerism; D010442:Peptide Chain Initiation, Translational; D012358:RNA, Transfer, Met / Q000378:metabolism
D000596:Amino Acids; D012358:RNA, Transfer, Met