SECONDARY STRUCTURE OF THE R(CUUCGG) TETRALOOP

The oligonucleotide r(GGACUUCGGUCC) has been observed to adopt a hairp in conformation by solution NMR and a double helical conformation by X -ray diffraction, In order to understand this apparent conflict, we us ed time-resolved fluorescence depolarization and (19)fluorine NMR to f ollow the secondary structure of this dodecamer as the solution compos ition was changed stepwise from the NMR experimental conditions to tho se used for crystallization. Calculation of the dodecamer concentratio n in the crystal (180 mM strands) and the cation concentration needed for neutrality (> 2 M) prompted investigation of a tethered species, i n which two dodecamers are connected by a string of 4 nt, geometricall y equivalent to similar to 100 mM strands, in 2.5 M NaCl, The RNA tetr aloop and its DNA analog maintain a single-strand hairpin conformation in solution, even under the conditions used to grow the crystal, Unde r high salt conditions, the tethered RNA and DNA analogs of this seque nce yield secondary components which could be the double helical confo rmation, Crystal contacts in addition to solvent changes and high RNA concentrations are needed to obtain the double helix as the predominan t species.