Fig 1.
Spectrophotometric titrations of (a) A 5 × 10−5 M GW-2974 with 10.0 μl increments of human telomeric G-quadruplex DNA (1.44 × 10−4 M). (b) A 4.00 × 10−6 M human telomeric G-quadruplex DNA with 10.0 μl increments of GW-2974 (1 × 10−5 M). (c) A 5 × 10−5 M SCH-442416 with 5.0 μl increments of human telomeric G-quadruplex DNA (1.44 × 10−4 M). All titrations were carried out in Tris–KCl buffer, pH 7.4.
Fig 2.
Fluorescence titrations of (a) A 5 × 10−6 M GW-2974 with human telomeric G-quadruplex DNA (1.33 × 10−4 M). (b) A 5 × 10−6 M of SCH-442416 with human telomeric G-quadruplex DNA (1.33 × 10−4 M). (c) A 2 × 10−6 M fluoresceine-labelled G-quadruplex DNA with 1 × 10−4 M GW-2974. (d) A 2 × 10−6 M fluorescein-labelled G-quadruplex DNA with 1 × 10−4 M SCH-442416. All titrations were carried out in Tris–KCl buffer, pH 7.4. Fluorescein-labelled G-quadruplex DNA had an excitation wavelength of 494 nm and emission wavelength of 518 nm.
Fig 3.
CD titration of human telomeric G-quadruplex DNA (1 × 10−5 M) with 1 × 10−4 M of GW-2974 (a) and SCH-442416 (b) in Tris–KCl buffer (pH 7.4).
Fig 4.
Stoichiometric ratio of [Drug]/{G-quadruplex DNA] using the molar ratio method.
Hunan telomeric G-quadruplex DNA (1.44 × 10−5 M) was added in different increments to 2.0 ml of 5 × 10−6 M GW-2974 (a) or SCH-442416 (b). Fluorescence emission of the two compounds was followed at 410 or 420 nm, respectively. Measurements were carried out in Tris–KCl buffer, pH 7.4.
Fig 5.
Increasing concentrations of human telomeric G-quadruplex DNA (1.44 x 10−4 M) were added to 2.0 ml of (5 × 10−6 M) GW-2974 (a, b) and SCH-442416 (c, d). The reaction was followed by measuring fluorescence emission of both compounds in Tris–KCl buffer, pH 7.4.
Table 1.
Binding constants (K) and number of binding sites (n) of GW-2974 and SCH-442416 per a G-quadruplex DNA molecule.
Fig 6.
Melting temperature curves for human telomeric G-quadruplex DNA and its complexes with (a) GW-2974 and (b) SCH-442416 using 1:1, 3:1 and 5:1 [(Drug)/(G-quadruplex)] molar ratios. Equimolar concentrations of 3.93 x 10−6 M were used. Fig 6c shows the melting temperature curves for ct-DNA and its complexes with GW-2974 and SCH-442416. Equimolar concentrations of 1 x 10−9 M were used.
Table 2.
Melting temperature of human telomeric G-quadruplex DNA and its GW-2974 and SCH-442416 complexes at different drug—DNA ratios.
ΔTm measures the difference between Tm’s of the drug-DNA complex and pure DNA.
Fig 7.
Selectivity of GW-2974 and SCH-442416 towards telomeric G-quadruplex DNA.
Fluorescence of fluorescein-labelled G-quadruplex DNA (5 × 10−10 M) complexed with equimolar GW-2974 and SCH-442416 were measured in the presences of 10.0, 50.0 and 100.0 folds of telomeric double-stranded DNA (a, c) and ct-DNA (b, d). Measurements were performed in Tris–KCl buffer, pH 7.4.
Table 3.
Docking scores of GW-2974, SCH-442416 and the co-crystallized drugs (quercetin and epiberberine) into the binding sites of the parallel (2MS6) and the hybrid (6CCW) human telomeric G-quadruplex DNA structures.
Fig 8.
a. Docking of GW-2974 and SCH-442416 into the binding sites of the parallel human telomeric G-quadruplex DNA structure (2MS6). Predicted binding modes of GW-2974 (pink sticks) and SCH-442416 (blue sticks) on G-quadruplex DNA (green sticks and cartoon). The 2D ligand interaction diagram shows the hydrogen bonding and π–π interaction as purple and green dotted lines, respectively. b. Docking of GW-2974 and SCH-442416 into the binding sites of the hybrid telomeric G-quadruplex DNA structure (6CCW). The G-quadruplex DNA with the two docked compounds is shown in (a). The 2D interaction diagrams show the binding modes of GW-2974 as pink sticks (a) and of SCH-442416 as green sticks in (b). c. The RMSD analysis of the docked complexes of the Telomeric G-quadruplex DNA with GW2974 (black), SCH-442416 (red), and the co-Crystallized ligand (green).