Equilibrium association constants for oligonucleotide-directed triple helix formation at single DNA sites: linkage to cation valence and concentration.
The linkage between the energetics of oligonucleotide-directed triple helix formation and the cationic solution environment has been investigated in mixed-valence salt solutions. Equilibrium constants for formation of the local pyrimidine.purine.pyrimidine structure afforded by binding of the oligonucleotide 5'-d(T*TTTTCTCTCTCTCT)-3' to a single site within a 339-bp plasmid fragment were measured using quantitative affinity cleavage titrations at pH 7.0 and 22 degrees C in the presence of various concentrations of KCl, MgCl2, and spermine tetrahydrochloride (SpmCl4). In a solution containing 10 mM NaCl, 140 mM KCl, 1.0 mM MgCl2, and 1.0 mM SpmCl4, the measured binding constant was 3.3 (+/- 1.4) x 10(5) M-1. The equilibrium constant previously reported for the same association reaction in 100 mM NaCl and 1 mM SpmCl4 at the same temperature and pH was 10-fold higher [Singleton, S. F., & Dervan, P. B. (1992) J. Am. Chem. Soc. 114, 6957-6965]. Further study demonstrated that varying the potassium ion concentration between 5.0 and 140 mM (in the presence of 10 mM NaCl, 1.0 mM MgCl2, and 1.0 mM SpmCl4) resulted in an overall 100-fold decrease in the binding affinity from the lowest to the highest concentration. In contrast, measured binding constants increased 500-fold as the spermine concentration was increased from 0.40 to 4.0 mM (in the presence of 10 mM NaCl, 140 mM KCl, and 1.0 mM MgCl2). There was a modest effect on the binding constant (a 3-fold decrease) upon varying the magnesium ion concentration from 0.10 to 10 mM (in the presence of 10 mM NaCl, 140 mM KCl, and 1.0 mM SpmCl4).(ABSTRACT TRUNCATED AT 250 WORDS)
Singleton
S F
SF
Arnold and Mabel Beckman Laboratories of Chemical Synthesis, California Institute of Technology, Pasadena 91125.
Dervan
P B
PB
D000595:Amino Acid Sequence; D002412:Cations / Q000737:chemistry; D004492:Edetic Acid / Q000737:chemistry; D006860:Hydrogen Bonding; D046911:Macromolecular Substances; D008274:Magnesium / Q000737:chemistry; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*; D011188:Potassium / Q000737:chemistry
D002412:Cations; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D004492:Edetic Acid; D008274:Magnesium; D011188:Potassium