Synthesis of azide congeners of preQ1 as potential substrates for tRNA guanine transglycosylase

PreQ(1) (2) is a precursor of queuine (1) that in eubacteria is incorporated into transfer RNA (tRNA) by tRNA guanine transglycosylase (TGT) before being further elaborated into queuine. The queuine modification is unusual and occurs across all eukaryotes and eubacteria with few exceptions, but its function remains unclear. As the modified nucleotide occurs through incorporation of a specially synthesized nucleotide instead of via modification of a genetically encoded base, a study of the sites of modification by prepared probes is possible. We report the synthesis of two novel azide congeners (3,4) of preQ(1) for this purpose. The evaluation of their interaction with TGT shows that both probes act as weak competitive inhibitors of guanine exchange of guanine(34) tRNA(Tyr) with a K-i of similar to 70 mu M. However, we could not show that these are substrates for TGT-catalyzed incorporation into tRNA. We believe the reason for this is a marked loss of binding due to the azide functionality of 3 and 4 abrogating the possibility of two hydrogen bonds to the carbonyl group of Leu231 and Met260 of TGT, previously observed for the terminal methylene amine of preQ(1) by x-ray crystallography.

Automated summary

PreQ(1) is a precursor of queuine that is incorporated into tRNA by TGT in bacteria and eukaryotes, but its exact function remains unclear. Two novel azide congeners of PreQ(1) were synthesized as probes, showing weak competitive inhibition of TGT but not as substrates for TGT-catalyzed incorporation into tRNA due to a loss of binding with TGT. Further research is needed to fully understand the mechanism.