Book of Abstracts: Albany 2007
June 19-23 2007
Structural Characterization of the Helix 31 Region of E. coli 16S rRNA and H. sapiens 18S rRNA Containing Different Post-transcriptional Modifications
E. coli 16S ribosomal RNA contains a total of eleven modified nucleotides. Most of them are located at or near the decoding region of the ribosome. The 970 loop region (U960-A975) of 16S rRNA is located near the ribosomal P-site and contains two modified nucleotides, m2G 966 and m5C 967. Structural and biological studies suggest that the 970 loop plays an important role in the decoding process as well as in tetracycline binding. To investigate the effects of these modifications, we have constructed helix 31, with and without modified nucleotides so that we can compare their effects using thermal melting, circular dichroism and NMR studies. The synthesis of the 6-O-DPC-2-N-methylguanosine (m2G) nucleoside, 5'-O-DMT-2'-O-TOM-protected 6-O-DPC-2-N-methylguanosine phosphoramidite and the biophysical data for the analogs of E. coli helix 31 will be presented [DPC, diphenyl carbamoyl; DMT, 4, 4'-dimethoxytrityl; TOM, [(triisopropylsilyl)oxy]methyl]. The two methylated bases, m2G966 and m5C967, occur at the same locus as the hypermodified nucleotide 1-methyl-3-(3-amino-3-carboxypropyl) pseudouridine (acp3m1Ψ) in H. sapiens. The syntheses of acp3m1Ψ and hence 5'-O-DMT-2'-O-TBS-protected acp3m1Ψ phosphoramidite will allow the selective incorporation of acp3m1Ψ into H. sapiens analogs. The availability of the phosphoramidites allow the selective incorporation and hence the syntheses of hairpin RNAs containing those modifications in the loop regions. These studies will reveal the roles of these different modifications on helix 31 structure and function.