# Ru(II)-Dppz Derivatives and Their Interactions with DNA: Thirty Years and Counting
## 3 DNA Binding
### Ru(II) Complexes as Intercalators  
In the last decades, more attention was given to metal complexes with an octahedral geometry, to verify if the species with high steric hindrance would intercalate and how this influences the process. In fact, ancillary ligands could prevent the insertion of small molecules between DNA adjacent base pairs. Ru(II) complexes containing rigid aromatic rings (stable, inert, and soluble in water) were among the first classes of octahedral complexes studied [61; 62]. One of these was [Ru(phen)\({}_{3}\)]\({}^{2+}\) species (phen = 1,10-phenanthroline) [63; 64; 65]. In this complex, each ligand has an aromatic planar structure that might interact with DNA; moreover, the three ligands give a chiral effect to the species. Chirality might have a relevant effect on the interaction process--according to [PERSON], the intercalation is favored for \(\Delta\) isomers of these kinds of complexes. This is due to the orientation of ligands in the \(\Delta\) isomers, which interfere sterically less than those in \(\Lambda\) isomers, with the skeleton of DNA. Later studies by [PERSON] and [PERSON] clarified that rather than intercalation, it is more correct to refer to this interaction as semi-intercalation, since there is only a partial insertion of a phenanthroline between the nucleobases, without the complete separation of the latter [10; 11; 12]. Taking enantioselectivity of Ru(II) tris-phenanthroline complexes as a starting point for designing probes that are able to recognize the double helix chirality, there were synthesized complexes that could selectively bind to DNA. For example, \(\Lambda\) isomers of metal complexes based on 4,7-diphenylphenanthroline (DIP) are efficient probes for Z-DNA [66; 67]. [Ru(TMP)\({}_{3}\)]\({}^{2+}\) (TMP = 3,4,7,8-tetramethylphenanthroline) is, instead, a selective probe for A-DNA [68]. Methyl substituents, in fact, do not allow the complex to intercalate between nucleobases; it interacts with DNA through electrostatic and hydrophobic interactions. This complex is too huge to insert into a B-DNA minor groove, but can insert into the major one of A-DNA and, through photoactivation and subsequent generation of \({}^{1}\)O\({}_{2}\), can break the double helix. However, the real research success in this field, and in fact, one of the most studied complexes is [Ru(\(\mathrm{bpy}\))\({}_{2}\)(dppz)]\({}^{2+}\) (dppz = dipyrido[3,2-a:2?,2?-c phenazine]); see Figure 1.  
The dppz ligand possesses a large aromatic surface area, allowing extensive intercalation in the DNA base pairs. Most Ru polypyridine complexes based on this ligand bind strongly to the DNA duplex via the intercalation mechanism. One measure of the affinity of these interactions is the log K value (K= DNA binding constant). For the classic DNA intercalator ethidium bromide (EB), this value is \(>\)6, while for [Ru(\(\mathrm{bpy}\))\({}_{2}\)dppz]\({}^{2+}\), under the same experimental conditions, this value is \(>\)7 [69], demonstrating the high affinity of this ligand for DNA. In 1990, [PERSON] first reported that [Ru(\(\mathrm{bpy}\))\({}_{2}\)(dppz)]\({}^{2+}\) could serve as a luminescent DNA probe [70]. This species does not exhibit luminescence in aqueous solution at room temperature, but shows intense emission in the presence of DNA, due to the radiative decay of an MLCT excited state, once the complex intercalates into the base pairs [71]. In water solution, instead, the luminescence is quenched due to proton-transfer from water molecules to phenazine's nitrogen atoms. Intercalation does not permit the interaction between water molecules and the Ru(II) complex, so the latter becomes luminescent. This phenomenon is known as the 'light-switch' effect of DNA and is widely used to study the interaction of polypyridyl metal complexes with DNA.