STUDIES ON FORMATION AND STABILITY OF THE (AG)(5)]ASTERISK-D[G(AG)(5)]CENTER-DOT-D[C(TC)(5)] AND (TG)(5)]ASTERISK-D[G(AG)(5)]CENTER-DOT-D[C(TC)(5)] TRIPLE HELICES

We have targeted the d[G(AG)(5)]. d[C(TC)(5)] duplex for triple format ion at neutral pH with either d[G(AG)(5)] or d[G(TG)(5)]. Using a comb ination of gel electrophoresis, uv and CD spectra, mixing and melting curves, along with DNase I digestion studies, we have investigated the stability of the 2:1 purpur . pyr tripler, d[G(AG)(5)]*d[G(AG)(5)]. d[C(TC)(5)], in the presence of MgCl2. This tripler melts in a monopha sic fashion at the same temperature eas the underlying duplex. Althoug h the uv spectrum changes little upon binding of the second purine sti -and, the CD spectrum shows significant changes in the wavelength rang e 200-230 nm and about a 7 nm shift in the positive band near 270 nm. In contrast, the 1:I:I pur/pyrpur . pyr triplex, d[G(TG)(5)]*d[G(AG)( 5)]. d[C(TC)(5)], is considerably less stable thermally, melting at a much lower temperature than the underlying duplex, and possesses a CD spectrum that is entirely: negative from 200 to 300 nm. Ethidium bromi de undergoes a strong fluorescence enhancement upon binding to each of these triplexes, and significantly stabilizes the pur/pyrpur . pyr t ripler. The uv melting and differential scanning calorimetry analysis of the alternating sequence duplex and purpur . pyr tripler shows tha t they ale lower in thermodynamic stability than the corresponding 10- mer d(G(3)A(4)G(3)). d(C3T4C3) duplex and its purpur . pyr triplex un der identical solution conditions. (C) 1997 John Wiley & Sons, Inc.