Abstract
A hexanucleotide GpGpGpUpGpG corresponding to bases 5-10 of the tRNAMetf from E. coli. has been synthesized by stepwise addition of mononucleotides. The segment contained five guanine bases out of six nucleotides. We have improved the protection of the 2', 3'-hydroxyl groups of guanosine and guanosine 3'-phosphate by replacing to avoid undesirable deblocking during elongation of the chain. Condensation of 5'-O-monomethoxytrityl-2'-O-benzoyl-N-isobutyrylguanosine 3'-phosphate (9) with 2', 3'-O-dibenzoyl-N-isobutyrylguansoine (5) gave protected GpG in a yield of 62%. Subsequent addition of mononucleotides to the growing chain was performed after selective removal of the 5'-monomethoxytrityl group. Dicyclohexylcarbodiimide (DCC) was used as condensing reagent throughout the synthesis. The excess of mononucleotide and the yield in each addition reaction were as follows : UpGpG (protected) 2.2 fold, 28%, GpUpGpG (protected), 5 fold, 32% ; GpGpUpGpG (protected) 10 fold, 35% ; GpGpUpGpG, 10 fold, 12%. The protected intermediate oligomers were isolated by ion-exchange chromatography on TEAE-cellulose columns and identified by enzymatic hydrolyses after deblocking. The hexanucleotide was separated by gel filtration on Sephadex LH-20 and purified on DEAE-cellulose in the presence of 7 M urea at 55°. The circular dichroism spectra of these guanine rich ribooligomers have measured and a marked difference between the pentamer GpGpUpGpG and the hexamer GpGpUpGpG has been observed.
