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Nucleic Acids Research logoLink to Nucleic Acids Research
. 1992 Oct 11;20(19):5159–5166. doi: 10.1093/nar/20.19.5159

Chemical synthesis of a biologically active natural tRNA with its minor bases.

D Gasparutto 1, T Livache 1, H Bazin 1, A M Duplaa 1, A Guy 1, A Khorlin 1, D Molko 1, A Roget 1, R Téoule 1
PMCID: PMC334300  PMID: 1383941

Abstract

The complete chemical synthesis of an E. coli tRNA(Ala) with its specific minor nucleosides, dihydrouridine, ribothymidine and pseudouridine, is reported. The method makes use of protected 2'-O-tertiobutyldimethylsilyl-ribonucleoside-3'-O-(2-cyanoethyl-N- ethyl-N- methyl)phosphoramidites. The exocyclic amino functions of the bases were protected by the phenoxyacetyl group for purines and acetyl for cytosine. The assembling has been performed on a silica support with coupling yield better than 98% within 2 min of condensation. Triethylamine tris-hydrofluoride allowed a clean and complete deprotection of the tBDMS groups. The synthetic tRNA(Ala) has been transcribed into cDNA by reverse transcriptase and sequenced. With E. coli alanyl-tRNA synthetase the alanyl acceptance activity and kcat/Km were 672 pmol/A260 and 6 x 10(4)M-1s-1, respectively.

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Selected References

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