Initiating translation with D-amino acids

Abstract

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.

Keywords

• D-amino acid
• translation
• initiation
• genetic code reprogramming
• flexizyme