STUDIES ON TRANSFER RIBONUCLEIC-ACIDS AND RELATED COMPOUNDS .29. SYNTHESIS OF A DECARBONUCLEOTIDE OF ESCHERICHIA-COLI TRANSFER-RNA-FMET (BASES 11-20) USING A NEW PHOSPHORYLATING REAGENT

A decanucleotide corresponding to the bases 11-20 of Escherichia coli tRNAfMet,3A-G-C-A-G-C-C-U-G-Gp4, was synthesized by triester block condensations. Completely protected mononucleotides (4) were prepared as the key intermediates by phosphorylation of the 2’-O-(o-nitrobenzyl) nucleosides (3) with a new phosphorylating reagent, p-chlorophenyl-phenyi-chlorophosphoramidate (2). The 5’-monomethoxytrityl group was removed and 5 were used as the 3’-terminal units in the synthesis of protected C-Up (9) and A-G-Cp (13). The diesterified mononucleotides (6) served as internal units. Successive coupling of these oligonucleotides to the dianilidated dinucleotide (8) yielded the protected decanucleotide (16). Mesitylenesulfonyl triazolide was used as the condensing reagent in both the synthesis of blocks and their condensation. The yields in condensations of monomers were 63-85% and those in block condensations were 27-60%. After deblocking of the terminal phosphate, amino, internucleotidic phosphate, and 5’-hydroxyl groups, the 2’-O-protected decanucleotide was irradiated with UV light to remove the o-nitrobenzyl ether. The deblocked decanucleotide was isolated by ion-exchange chromatography on DEAE-cellulose and characterized by complete hydrolysis with a base-nonspecific RNase to give a correct ratio of mononucleotides. The sequence was confirmed by mobility shift analysis. The decanucleotide is the largest naturally occurring fragment synthesized chemically. © 1979, American Chemical Society. All rights reserved.