A n approach to the solid-phase synthesis of oligo-and poly-ribonucleotides is described. The synthetic strategy involves the use of building blocks in which t w o acid-labile groups, 1 -(2-fluorophenyl) -4-methoxypiperidin-4-yl (Fpmp) and 9-phenylxanthen-9-yl (Px), respectively, are used to protect the 2'and 5'hydroxy functions of ribonucleoside building blocks. The adenine, cytosine and guanine base residues are protected with pivaloyl, benzoyl and phenylacetyl groups, respectively. 2-Cyanoethyl N,N-diisopropylphosphoramidites are used in the coupling steps, and 5-(3nitrophenyl) -1 H-tetrazole is used as the activating agent. Following the chain-assembly process, 2'protected oligo-and poly-ribonucleotides are released from the functionalized controlled-pore glass solid support; the latter stabilized ribonucleic acid (RNA) sequences are purified before they are fully unblocked by treatment with 0.01 mol dms hydrochloric acid (pH 2) at room temperature for 20 h. The efficacy of this methodology is illustrated by the synthesis of the 3'-terminal decamer (r[ UCG UCCACCA] ), nonadecarner (r[AU UCCGGACUCG UCCACCA] ), and heptatriacontamer (37mer, r [ GGAGAGG UCUCCGG U UCGAU UCCGGACUCG UCCACCA] ) sequences of yeast alanine tRNA (tRNAAIa).

The preparation of a 4-thiouridine phosphoramidite suitable for RNA synthesis and its subsequent incorporation into oligoribonucleotides is described. The thiol group is protected with a 2-cyanoethyl group and the 2'-OH with a 1-(2-fluorophenyl)-4-methoxypiperidin-4-yl function. Thiouridine-containing oligoribonucleotides were used as 350 nm UV crosslinking probes for the photoaffinity labelling of RNA binding proteins. Specific crosslinking was demonstrated between the Rev protein of HIV-1 (as a glutathione S-transferase fusion protein) and its RNA target, the Rev-responsive element. It was not possible to generate crosslinks between the RNA bacteriophage MS2 coat protein and the initiator stem-loop of the replicase gene, to which it binds. These results are consistent with the structural data available on both systems.

No abstract

The synthesis of cyclic di-, tri-, tetra-, penta- and hexa-thymidylic acids (16; n = 2, 3, 4, 5 and 6) and the cyclic hexadeoxyribonucleotide [d(CpCpTpApGpGp)], via the "filtration" approach, is reported. Some of the physical properties of the cyclic oligonucleotides are discussed, and their susceptibility to digestion in the presence of phosphorylytic enzymes has been studied.

No abstract