Modulation of Cm/T, G/A, and G/T triplex stability by conjugate groups in the presence and absence of KCl.
Apparent equilibrium association constants were determined by gel mobility shift analysis for triple strand formation between a duplex target containing a 21 base long A-rich homopurine run and several end-modified C(m)/T (pyrimidine motif; C(m) = 5-methylcytosine), G/A (purine motif), and G/T (purine-pyrimidine motif) triplex-forming oligonucleotides (TFOs). Incubations were carried out for 24 h at 37 degrees C in 20 mM HEPES, pH 7.2, 10 mM MgCl2, and 1 mM spermine. The purine motif triplex was the most stable (Ka = 6.2 x 10(8) M-1) even though the TFO self-associated as a linear duplex. Conjugation of a terminal hexanol or cholesterol group to the G/A-containing TFO reduced triplex stability by 1.6- or 13-fold, whereas an aminohexyl group or intercalating agent (acridine or psoralen) increased triplex stability by 1.3- or 13-fold. These end groups produced similar effects in C(m)/T and G/T triplexes, although the magnitude of the effect sometimes differed. Addition of 140 mM KCl to mimic physiological conditions decreased stability of the G/A triplex by 1900-fold, making it less stable than the C(m)/T triplex. The inhibitory effect of KCl on G/A triplex formation could be partially compensated for by conjugating the TFO to an intercalating agent (30-350-fold stabilization) or by adding the triplex selective intercalator coralyne (1000-fold stabilization). Although the G/T triplex responded similarly to these agents, the stability of the C(m)/T triplex was unaffected by the presence of coralyne and was only enhanced 1.4-2.8-fold when the TFO was linked to an intercalating agent. In physiological buffer supplemented with 40 microM coralyne, the G/A triplex (Ka = 3.0 x 10(8) M-1) was more stable than the C(m)/T and G/T triplexes by factors of 300 and 12, respectively.
Gamper
H B
HB
Epoch Pharmaceuticals, Inc., Bothell, Washington 98021, USA.
Kutyavin
I V
IV
Rhinehart
R L
RL
Lokhov
S G
SG
Reed
M W
MW
Meyer
R B
RB
D001482:Base Composition; D004247:DNA / Q000737:chemistry / Q000187:drug effects*; D006683:HLA-DQ Antigens / Q000235:genetics*; D059866:HLA-DQ beta-Chains; D006801:Humans; D009690:Nucleic Acid Conformation / Q000187:drug effects*; D009691:Nucleic Acid Denaturation / Q000187:drug effects; D009693:Nucleic Acid Hybridization / Q000187:drug effects; D009838:Oligodeoxyribonucleotides; D011189:Potassium Chloride / Q000494:pharmacology*
D006683:HLA-DQ Antigens; D059866:HLA-DQ beta-Chains; C069051:HLA-DQB1 antigen; D009838:Oligodeoxyribonucleotides; C051802:triplex DNA; D011189:Potassium Chloride; D004247:DNA