PREPARATION OF OLIGORIBONUCLEOTIDES CONTAINING 4-THIOURIDINE USING FPMP CHEMISTRY - PHOTO-CROSS-LINKING TO RNA-BINDING PROTEINS USING 350 NM IRRADIATION

The preparation of a 4-thiouridine phosphoramidite suitable for RNA sy nthesis 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. T hiouridine-containing oligoribonucleotides were used as 350 nm UV cros slinking probes for the photoaffinity labelling of RNA binding protein s. Specific crosslinking was demonstrated between the Rev protein of H IV-1 (as a glutathione S-transferase fusion protein) and its RNA targe t, the Rev-responsive element. It was not possible to generate crossli nks between the RNA bacteriophage MS2 coat protein and the initiator s tem-loop of the replicase gene, to which it binds, These results are c onsistent with the structural data available on both systems.