**3. Native G-quartet-based DNAzymes: G-quadruplexes**

## **3.1. Structure of G-quadruplexes**

G-quadruplexes (Figure 1), formed from G-rich DNA strands, are composed of a stacking of several native G-quartets. Each quartet results from the self-assembly of four guanines in a same plan, self-stabilized by eight hydrogen bonds, from the Hoogsteen and Watson–Crick faces of the guanine moieties. The additional π-stacking interactions between G-quartets, and the bonding of cationic ions (*e.g*., K+ , Na+ ), increase the global stability of the tridimensional edifice.[10, 26, 27] In a structural point of view, the G-quadruplexes can mainly differ from the total numbers of constitutive G-quartets (from two to several thousands)[28–30] or strands (from one to four),[31–35] from the orientation of the strands (leading to several conformations named antiparallel, parallel, and hybrid), and finally from the length, DNA bases composition, and position of the loop(s) (which can be edgewise, diagonal, or chain-reverse).[36–41] All of these parameters are linked to the global stability of the edifice, like the number of G-quartets (as it is discussed later in this chapter), and are interdependent.

**Figure 1.** Schematic representation of the self-assembly of guanines *via* the Watson–Crick and Hoogsteen faces to form G-quartets, and G-quadruplexes.

These noncanonical structures are well known in a biological context, because they are strongly suspected to play important roles in key cellular events, like chromosomal instability, or regulation of gene expression. These aspects are far from the scope of this chapter, and author incites curious readers to have a look to some reviews cited hereafter.[26, 27, 42–44]

## **3.2. The seeds of the G-quartet ability to catalyze peroxidase-like reactions**

In 1996, Y. Li and D. Sen developed and published the fourth known DNAzyme system,[45] able to catalyze the incorporation of metals (*i.e.*, Cu(II) and Zn(II)) into a specific porphyrin, named mesoporphyrin IX, or MPIX. To select the best DNA catalyst for their system, they used the *in vitro* SELEX (for *systematic evolution of ligands by exponential enrichment*) method that highlighted one sequence, termed PS5.ST1, from an initial pool of DNA sequences.[45, 46] Interestingly, three main observations were crucial:


The subsequent step has been taken in 1998 when the same group showed that hemin, a Fe(III) cofactor playing a pivotal role in many enzymes (*e.g*., in catalases, monooxygenases, and peroxidases), is activated by the presence of a G-quadruplex. More precisely, authors proved that the activity of the *horseradish peroxidase* (also termed HRP), composed of an hemin surrounded by a protein environment, can be mimicked using the same hemin but in the presence of G-quadruplexes.[50] Since then, many experiments using different sequences and morphologies of G-quadruplexes unambiguously confirmed the mandatory role of the Gquadruplex structure, for which the accessible G-quartet (*i.e*., the external one), able to interact with hemin, constitutes the key step of the reaction.[24, 51–55]
