Poster abstracts

Poster number 16 submitted by Akwasi Agyeman

The effect of conserved sequence variation on the binding of the antiterminator element to tRNA

Akwasi Agyeman (Chemistry and Biochemistry, Ohio University), Hamid Fauzi (Chemistry and Biochemistry, Ohio University), Abigael Muchenditsi (Chemistry and Biochemistry, Ohio University), Jennifer Hines (Chemistry and Biochemistry, Ohio University)

Abstract:
A critical interaction that occurs during the T box transcriptional antitermination regulatory mechanism involves a unique RNA-RNA interaction. The acceptor end of an uncharged cognate tRNA interacts with 5'-end nucleotides in a 7 - nucleotide bulge of an otherwise unstable antiterminator element to stabilize it. The nucleotides that form the bulge of the antiterminator are the middle 7 sequence of the 14-nucleotide T box sequence, which is highly conserved. The significance of the conservation of the 3'-end nucleotides of the bulge and in the A2 helix above the bulge, though unknown, has been found to be very important for function. To determine whether the antiterminator and tRNA requirements for functional antitermination are determined solely at the level of RNA-RNA interactions, a series of antiterminator and tRNA model RNA have been investigated. The antiterminator RNAs were selected based on phylogenetic-derived sequence changes and bulge sequences derived from in vitro evolution studies. Affinities and general structural characteristics between the antiterminator models and tRNA analogs were investigated using fluorescence anisotropy, enzymatic probing and circular dichroism. Our results show that there are structural differences between the antiterminator RNAs and that they bind the tRNA analogs with differing affinities. Although there were some exceptions, our results correlate very well with in vivo and in vitro antitermination efficiency data. The implications of the findings from this study will be discussed in detail.

Keywords: Antiterminator, Phylogenetic, Anisotropy