Characterization of the DNA triplex formed by d(TGGGTGGGTGGTTGGGTGGG) and a critical R.Y sequence located in the promoter of the murine Ki-ras proto-oncogene.
The binding of the G-rich oligonucleotide d(TGGGTGGGTGGTTGGGTGGG) to a critical homopurine-homopyrimidine sequence located in the promoter of the murine Ki-ras proto-oncogene has been investigated. The duplex and the oligonucleotide form a triple helix as evidenced by band-shift electrophoresis, hydroxyapatite (HA) chromatography, UV-melting and circular dichroism (CD) experiments. Upon thermal denaturation in 50 mM Tris-acetate, pH 7.4, 50 mM NaCl, 10 mM MgCl2, 0.1 mM spermine the triplex exhibits two cooperative transitions: one of these is attributed to the triplex-to-duplex transformation, the other to the duplex-to-coil transformation. The thermodynamic parameters of triplex formation have been determined by a van't Hoff analysis of the UV-melting curves which provided values of delta H = 79 +/- 8 kcal/mol, delta S = 224 +/- 22 e.u., delta G298 = 12.2 +/- 1.2 kcal/mol. These data are compared with those reported for the YRY triplex motif.
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Department of Biochemistry, Biophysics and Macromolecular Chemistry, University of Trieste, Italy.
D000818:Animals; D001483:Base Sequence; D002151:Calorimetry; D002942:Circular Dichroism; D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D011905:Genes, ras; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification; D011401:Promoter Regions, Genetic; D013816:Thermodynamics
D009838:Oligodeoxyribonucleotides; D004247:DNA