The Secondary Structure of a Fourteen-Nucleotide Fragment of the Hairpin Ribozyme

Abstract

A fourteen nucleotide RNA has been synthesized and its secondary structure investigated by non-denaturing polyacrylamide gel electrophoresis and1H nuclear magnetic resonance spectroscopy. This fourteen nucleotide RNA corresponds to the hairpin loop end of the proposed secondary structure of the (−)sTRSV hairpin ribozyme. Non-denaturing polyacrylamide gel electrophoresis indicates that the fourteen nucleotide RNA exists predominantly as a monomer at a 1 mM strand concentration. Four peaks are found in the imino hydrogen region of the1H NMR spectrum of the fourteen nucleotide RNA at this concentration. One-dimensional nuclear Overhauser effect spectroscopy of the imino hydrogen region of the1H NMR spectrum gives results consistent with a model of the secondary structure of the fourteen nucleotide RNA having a three nucleotide hairpin loop and two double-stranded stems separated by a single bulged adenosine.