Chemical biology studies of an RNA transcription molecular switch: Sequence and metal ion effects on T box antiterminator RNA•tRNA binding

BIOL 96

Jennifer V. Hines, hinesj@ohio.edu, Akwasi Agyeman, Rajaneesh Anupam, ra357002@ohio.edu, Hamid Fauzi, Karen D. Jack, John A. Means, john.a.means.1@ohio.edu, Abigael M. Muchenditsi, and Crystal Simson. Department of Chemistry and Biochemistry, Ohio University, Clippinger Laboratories, Athens, OH 45701
The transcription of Gram-positive bacterial genes in the T box family depends on the stabilization of an antiterminator element in the nascent mRNA transcript. The stabilization occurs by base pairing of uncharged cognate tRNA with the antiterminator and precludes the formation of a competing terminator element. In this manner, uncharged tRNA serves as the molecular effector for this transcriptional RNA switch enabling the bacteria to carefully regulate amino acid related metabolic genes. To determine whether the antiterminator and tRNA sequence requirements for functional antitermination are determined solely at the level of RNA-RNA interactions and to investigate the role divalent metal ions play in this molecular recognition, a series of antiterminator and tRNA model RNAs have been investigated. Affinities and general structural characteristics were investigated using fluorescence, UV, CD and NMR spectroscopy along with enzymatic probing. The significance of the sequence and metal ion effects controlling functionally-relevant binding will be presented.