Differential DNA Recognition by the Enantiomers of 1-Rh(MGP)_2phi: A Combination of Shape Selection and Direct Readout

The enantiomers of the symmetric metallointercalator complex 1-Rh(MGP)_2phi^(5+) [MGP = 4-(guanidylmethyl)-1,10-phenanthroline; phi = phenanthrenequinone diimine] bound to DNA decamer duplexes containing their respective 6 bp recognition sequences have been investigated using ^1H NMR. Shape selection due to the chirality of the metal center and hydrogen-bonding contacts of ancillary guanidinium groups to 3‘-G N7 atoms define the recognition by complexes which bind by intercalation to duplex DNA. The titration of Λ-Rh into the self-complementary decamer containing the recognition sequence (5‘-GACATATGTC-3‘, L1) resulted in one symmetric bound conformation observed in the ^1H NMR spectrum, indicating that the DNA duplex retains its symmetry in the presence of the metal complex. Upfield chemical shifts of duplex imino protons and the disruption of the NOE base−sugar contacts defined the central T5-A6 intercalation site. The downfield shift of the G8 imino proton supports the conclusion that the pendant guanidinium arms make simultaneous H-bonding contacts to the N7 atoms of 3‘-G8 bases on either side of the site. A variable-temperature study of a partially titrated sample (2:3 Λ-Rh/L1) showed the exchange rate (k_(obs)) at 298 K to be 68 s^(-1) and the activation barrier to exchange (ΔG^⧧ of association) to be 2.7 kcal/mol, a value comparable to the stacking energy of one base step. The results presented coupled with biochemical data are therefore consistent with binding models in which Λ-1-Rh(MGP)_2phi^(5+) (Λ-Rh) traps the recognition site 5‘-CATATG-3‘ in an unwound state, permitting intercalation centrally and hydrogen bonding to guanines at the first and sixth base pair positions. The data suggest a different model of binding and recognition by Δ-Rh. The titration of Δ-Rh into a DNA decamer containing the 6 bp recognition site (D1, 5‘-CGCATCTGAC-3‘; D2, 5‘-GTCAGATGCG-3‘) resulted in two, distinct conformers, in slow exchange on the NMR time scale. The rate of exchange between the two conformers (k_(obs)) at 298 K is 37 s^(-1), most likely due to partial dissociation between binding modes. The slower rate relative to Λ-Rh association reflects the relative rigidity of the D1 and/or D2 sequence in comparison to L1. NOE cross-peaks between the intercalating phi ligand and protons of T5-C6, as well as the upfield shifts observed for imino protons at this step, serve to define the central T5-C6 step as the single site of intercalation. The downfield shift of the 3‘-G imino protons indicates the complex makes hydrogen bond contacts with these bases. The complex, which is too small to span a 6 bp B-form DNA sequence, nonetheless makes major groove contacts with 3‘-G bases to either side of the site. Notably, both 3‘-guanine bases are necessary to impart site specificity and slow dissociation kinetics with the 5‘-CATCTG-3‘ site, as evidenced by the extremely exchange-broadened two-dimensional NOESY spectra of Δ-Rh bound to modified duplexes containing N7-deazaguanine at either G8 or G18; the loss of one major groove contact completely abolishes specificity for 5‘-CATCTG-3‘. DNA chemical shifts upon binding and intermolecular NOE contacts therefore support a model in which Δ-Rh intercalates in one of two canted binding conformations. Within this model, each intercalation mode allows one guanidinium−guanine hydrogen bond at a time, while bringing the other arm close to the phosphate backbone