Synthesis of tRNA analogues containing a terminal ribose locked in the South conformation to study tRNA-dependent enzymes

Abstract

We report here the synthetic route of two constrained dinucleotides and the determination of the sugar puckering by NMR analyses of the starting nucleosides. Enzymatic ligation to microhelix-RNAs provide access to tRNA analogues containing a 3' terminal A⁷⁶ locked in South conformation. Biological evaluation of our tRNA analogues has been performed using amino-acyl tRNA-dependent transferase FemX_Wv, which mediates non-ribosomal incorporation of amino acids into the bacterial cell wall. We have shown that our tRNA analogues inhibited the aminoacyl transfer reaction catalyzed by FemX_Wv with IC_50s of 10 and 8 μM. These results indicate that FemX_Wv displays a moderate preference for tRNAs containing a terminal A⁷⁶ locked in the South conformation and that a South to North switch in the conformation of the terminal ribose might contribute to the release of the uncharged tRNA^Ala product of the aminoacyl transfer reaction catalyzed by FemX_wv.