Synthesis of 3'-fluoro-tRNA analogues for exploring non-ribosomal peptide synthesis in bacteria.

Aminoacyl-tRNAs (aa-tRNAs) participate in a vast repertoire of metabolic pathways, including the synthesis of the peptidoglycan network in the cell walls of bacterial pathogens. Synthesis of aminoacyl-tRNA analogues is critical for further understanding the mechanisms of these reactions. Here we report the semi-synthesis of 3'-fluoro analogues of Ala-tRNA(Ala) . The presence of fluorine in the 3'-position blocks Ala at the 2'-position by preventing spontaneous migration of the residue between positions 2' and 3'. NMR analyses showed that substitution of the 3'-hydroxy group by fluorine in the ribo configuration favours the S-type conformation of the furanose ring of terminal adenosine A76. In contrast, the N-type conformation is favoured by the presence of fluorine in the xylo configuration. Thus, introduction of fluorine in the ribo and xylo configurations affects the conformation of the furanose ring in reciprocal ways. These compounds should provide insight into substrate recognition by Fem transferases and the Ala-tRNA synthetases.

Synthesis of 3'-triazoyl-dinucleotides as precursors of stable Phe-tRNA(Phe) and Leu-tRNA(Leu) analogues.

We report here the synthesis of stable Phe-tRNA(Phe) and Leu-tRNA(Leu) analogues containing a 1,2,3-triazole ring instead of the ribose-amino acid ester bond. The 1,2,3-triazole ring is generated by dipolar cycloaddition of alkyne Phe and Leu analogues to 3'-azido-3'-deoxyadenosine via the Cu(I)-catalysed Huisgen, Meldal, Sharpless 1,3-cycloaddition. The corresponding triazoyl pdCpA dinucleotides, obtained by classical phosphoramidite chemistry, were enzymatically ligated to 22-nt or 74-nt RNA generating stable Phe-tRNA(Phe) analogues containing the acceptor stem or full tRNA moieties, respectively. These molecules represent useful tools to study the contribution of the RNA and amino acid moieties in stabilization of aminoacyl-tRNA/protein complexes.

Enhanced photoinduced electron transfer at the surface of charged lipid bilayers.

Photocatalytic systems often suffer from poor quantum yields due to fast charge recombination: The energy-wasting annihilation of the photochemically created charge-separated state. In this report, we show that the efficiency of photoinduced electron transfer from a sacrificial electron donor to positively charged methyl viologen, or to negatively charged 5,5'-dithiobis(2-nitrobenzoate), increases dramatically upon addition of charged phospholipid vesicles if the charge of the lipids is of the same sign as that of the electron acceptor. Centrifugation and UV/Vis titration experiments showed that the charged photosensitizers adsorb at the liposome surface, that is, where the photocatalytic reaction takes place. The increased photoelectron transfer efficiency in the presence of charged liposomes has been ascribed to preferential electrostatic interactions between the photosensitizer and the membrane, which prevents the formation of photosensitizer-electron-acceptor complexes that are inactive towards photoreduction. Furthermore, it is shown that the addition of liposomes results in a decrease in photoproduct inhibition, which is caused by repulsion of the reduced electron acceptor by the photocatalytic site. Thus, liposomes can be used as a support to perform efficient photocatalysis; the charged photoproducts are pushed away from the liposomes and represent "soluble electrons" that can be physically separated from the place where they were generated.