Triple helixes containing one homopurine poly dG or poly rG strand and two homopyrimidine poly dC or poly rC strands have been prepared and studied by FTIR spectroscopy in H2O and D2O solutions. The spectra are discussed by comparison with those of the corresponding third strands (auto associated or not) and of double stranded poly dG.poly dC and poly rG.poly rC in the same concentration range and salt conditions. The triplex formation is characterized by the study of the base-base interactions reflected by changes in the spectral domain involving the in-plane double bond vibrations of the bases. Modifications of the initial duplex conformation (A family form for poly rG.poly rC, B family form for poly dG.poly dC) when the triplex is formed have been investigated. Two spectral domains (950-800 and 1450-1350 cm-1) containing absorption bands markers of the N and S type sugar geometries have been extensively studied. The spectra of the triplexes prepared starting with a double helix containing only riboses (poly rC+.poly rG.poly rC and poly dC+.poly rG.poly rC) as well as that of poly rC+.poly dG.poly dC present exclusively markers of the North type geometry of the sugars. On the contrary in the case of the poly dC+.poly dG.poly dC triplex both N and S type sugars are shown to coexist. The FTIR spectra allow us to propose that in this case the sugars of the purine (poly dG) strand adopt the S type geometry.
We have studied the effect of the nature of the third-strand sugar (ribose or deoxyribose) on the geometry and stability of triple helices with a pyrimidine motif targeting the polypurine tract of the Friend murine retrovirus. Comparison between triplexes containing a third strand formed by a deoxy 13mer d(TCT5C6), the same oligomer but with C5-methylated cytosines d(T5meCT5(5me)C6), and an analogous modified 13mer RNA 2'Omer(UCU5C6) shows that the sugar conformations of the different triple helices, determined by FTIR spectroscopy, differ depending on nature of the third-strand sugar. Pyrimidine*purine-pyrimidine triple-helix formation with the third-strand RNA and the duplex as DNA appears to be associated with a conversion of the duplex part from a B-form secondary structure with S-type sugars to a geometry in which the polypurine strand sugars adopt an N-type conformation. Thermal dissociation of the triplexes was studied by UV absorbance spectroscopy. The most stable triple helix is obtained when the third strand contains 2'-O-methylated ribose sugars.
Many early investigations on triple helices have been devoted to the study of the triplex formed by dT*dA-dT base triplets in which the third strand is oriented parallel to the dA strand. We now describe an intramolecular triple helix with dT*dA-dT base triplets in which the pyrimidine third strand is oriented antiparallel, formed by folding back twice the tridecamer dT10-linker-dA10-linker-dT10 (linker = pO(CH2CH2O)3p). Third-strand base pairing to the target strand, sugar conformation, and thermal denaturation of the triplex have been studied by Fourier transform infrared spectroscopy. Our results confirm than when the third-strand orientation is reversed from parallel to antiparallel with respect to the target strand, the third-strand hydrogen-bonding scheme is changed from Hoogsteen to reverse Hoogsteen. The sugar conformation in this triple helix is of the S type (C2'endo/anti, B family form) from all strands. Our results are discussed with respect to models for triplexes proposed as intermediates in homologous recombination [Zhurkin, V.B., Raghunathan, G., Ulyanov, N.B., Camerini-Otero, R.D., & Jernigan, R.L. (1994) J. Mol. Biol. 239, 181-200].
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