Vibrational CD study of the thermal denaturation of poly(rA) · poly(rU)

Abstract

The vibrational cd (VCD) of a double-stranded RNA, poly(rA).poly(rU), at pH 7 and moderate added salt concentration (0.1 M) has been measured in both the base-stretching and phosphate-stretching regions of the ir as a function of temperature. The data in both cases show two distinct phase transitions. The first is from double- to a triple-stranded form, and the second is from triple- to single-stranded forms, which still retain substantial local order even up to 80 degrees C. The nature of these transitions has been identified by comparison of the VCD and ir absorption spectra of the initially double-stranded samples with those of single-stranded poly(rA) and poly(rU) and with triple-stranded poly-(rA).poly-(rU).poly(rU). The large differences in the VCD band shapes allows positive identification of the intermediate and final states. Thus under VCD-concentration conditions, a simple helix-to-coil transition can be eliminated for poly(rA).poly(rU) while such a two-step transition can be seen at low salt conditions. All of these observations are consistent with previous studies of the phase transitions of poly(rA).poly(rU) under various salt conditions. Additionally, the VCD is indicative of premelting for all the triple-, double-, and single-strand complexes studied. The triple-strand complex did not show disproportionation to double strand on heating under these added salt conditions. The unusual VCD pattern for low temperature poly(rA).poly(rU), as compared to high G--C content RNAs and DNAs, is qualitatively, but not quantitatively, explained using exciton coupling of localized dipolar transitions in each type of base within the strand.