Specific and nonspecific bindings of alkaline-earth metal ions to guanine-quadruplex thrombin-binding aptamer DNA

Specific and nonspecific bindings of alkaline-earth metal ions to the thrombin-binding aptamer (TBA) DNA d(GGTTGGTGTGGTTGG) were studied using electrospray ionization (ESI) mass spectrometry (MS). A single-stranded DNA d(AATTAATGTAATTAA) was used as a control for nonspecific binding. Both 1:1 and 2:1 metal–DNA complexes formed in 3:1 water/isopropanol were detected with ESI-MS and their binding constants were determined by the titration method. The 1:1 binding constant (K1) of Sr2+ or Ba2+ for TBA was three orders of magnitude greater than their K1 for the control DNA, whereas K1 of Mg2+ or Ca2+ for TBA was comparable to their K1 for the control DNA. To the contrary, the second binding constant (K2) was nearly the same independent of the metal ion and DNA. To see if addition of metal ion led to a structural change in DNA, the circular dichroism (CD) spectra of DNA in ESI solvent were obtained in the presence and absence of metal ion. TBA showed characteristic bands of the chair-type guanine-quadruplex (G4) structure. Significantly, addition of Sr2+ or Ba2+ enhanced the G4 bands of TBA, but neither Mg2+ nor Ca2+ induced a change, indicating that Sr2+ and Ba2+ interact strongly with G4 TBA and stabilize it. Control DNA showed no special motif and none of the four metal ions affected its CD spectra. Binding constants and CD results suggest that only the first binding of Sr2+ and Ba2+ to G4 TBA is specific and the first binding of Mg2+ and Ca2+ as well as the second binding of all four metal ions are nonspecific. The specific and nonspecific bindings of alkaline-earth metal ions to G4 TBA are further rationalized in terms of ionic radius, coordination number, and Gibbs free energy of dehydration of the metal ion.

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► The first bindings of Sr2+ and Ba2+ to G4 TBA are specific, whereas those of Mg2+ and Ca2+ are not.
► Sr2+ and Ba2+ specifically bind to the intercalation site of G4 TBA, thereby greatly stabilizing the G4 structure.
► Mg2+ and Ca2+ nonspecifically bind to the outer region of G4 TBA.
► The second bindings of all alkaline-earth metal ions to G4 TBA are nonspecific.