One of the long-term objectives in our group research is to understand the roles of the modified nucleotides present in ribosomal RNA. My research project involved the use of synthetic, biochemical and biophysical approaches to help us better understand the roles of methylated analogs. A major goal was to reveal the ability to utilize a functionally important region in the bacterial ribosome as a possible drug target. Primarily, my research focused on the characterization of the effects of modified nucleotides present in helix 31 (h31) of 16S ribosomal RNA. In particular, I synthesized the 6- O -DPC-2- N -methylguanosine (m 2 G) nucleoside and corresponding 5'- O -DMT-2'- O -TOM-protected 6- O -DPC-2- N -methylguano-sine phosphoramidite [DPC, diphenyl carbamoyl; DMT, 4, 4'-dimethoxytrityl; TOM, [(triisopropylsilyl)oxy]methyl]. Availability of the amidite allowed for incorporation of m 2 G into specific RNA regions of interest. The two methylated bases, m 2 G966 and m 5 C967, occur at the same locus as the hypermodified nucleotide 1-methyl-3-(3-amino-3-carboxypropyl) pseudouridine (acp 3 m 1 Ψ) in H. sapiens . I explored the significance of the E.coli modifications in h31 by using various biophysical techniques. My ultimate goal was to discover peptide ligand/s that can bind specifically to this region of E. coli 16S rRNA, and utilize helix 31 as a potential drug target in the bacterial ribosome. The availability of the wild-type h31 RNA construct allowed us to achieve this objective.
