Nucleic Acids Research, 1981, Vol. 9, No. 14 3503-3522
© 1981
CHEMISTRY |
Studies on transfer ribonucleic acids and related compounds. XL.1 Synthesis of an eicosaribonucleotide corresponding to residues 35–54 of tRNAfMet from E. coli
Faculty of Pharmaceutical Sciences, Osaka University Suita, Osaka 565, Japan
Received May 18, 1981.
An E.coli tRNAfMet fragment [C-A-U-A-A-C-C-C-G-A-A-G-G-U-CG-U-C-G-G (bases 35–54)] containing the anticodon triplet has been synthesized by the phosphotriester method involving protected oligonucleotide blocks. Di- or tri-nucleotide blocks were prepared by condensation of 2'-O-(o-nitrobenzyl)nucleotide derivatives and used for the synthesis of pentanucleotide blocks. The 5'-hydroxy, heterocyclic amino and internucleotide linkage were protected with monomethoxytrityl, acyl and p-chlorophenyl groups, respectively. The 3'-phosphates of the pentanucleotides, except for the GUCGG block where 2'-0-benzoyl 3'-O-(o-nitrobenzyl) N-isobutyrylguanosine was used, were protected with pchlorophenyl and anilido groups. The anilido groups were removed by treatment with isoamyl nitrite and the 3'-phosphodiesters of resulting pentamers were activated with mesitylenesulfonyl nitrotriazolide to give protected decanucleotides in yields of 61–89%. The two decanucleotides were condensed similarly to yield the protected eicosanucleotide in a yield of 59%. The product was deblocked and purified by ion-exchange chromatography on DEAE-Sephadex A-25 and characterized by enzymatic hydrolysis after labelling the 5'-end by phosphorylation using polynucleotide kinase and [
-32p]ATP.