To solve details of the self-assembling process of guanosine in diluted aqueous solution and to derive a thermodynamical model for quadruplex formation, the structural behavior of deoxyguanosine 5'-monophosphate has been analyzed by in-solution small angle X-ray scattering. The experiments have been performed as a function of guanosine concentration and at fixed guanosine concentration but in the presence of varying amounts of KCl. As a result, the self-assembling process, in terms of both aggregate particle fractions and aggregate length, has been observed to be strongly dependent on composition and largely affected by excess potassium ions in the solution. In particular, the different aggregate forms have been resolved and their concentration derived as a function of sample composition. In accordance with a hierarchical aggregation process, a nucleation and elongation mechanism has been used to derive the thermodynamical parameters for self-assembling. The results show that the annealing and fragmentation steps play an important role in the aggregation process.

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Guanosine, one of the primary components of nucleic acids, self-associates in water to form G-quadruplexes, four-stranded helicoidal aggregates, made by stacked planar tetramers, consisting of four planar guanine molecules. Essential for the stability of these supramolecular aggregates is the presence of monovalent cations. As G-quadruplexes show a lyotropic polymorphism, neutron diffraction, in combination with the H2O/D2O contrast variation technique, has been applied to study the cation structural features of quadruplexes in hexagonal phase at different hydrations and counterion concentrations. The guanosine 5'-monophosphate, dipotassium salt, was considered and G-quadruplexes in hexagonal phase were prepared in the different experimental conditions (contrast, hydration and KCl solution concentration) by using the osmotic stress technique. The calculated scattering length density distribution maps show that counterions fill the helix inner cavity and that atmospheric cations are bound to a second site, close to the external phosphate groups. The site occupancy turned out to be very high: we found on the inner site 0.87 K ions per tetramer in G-quadruplexes prepared in pure water and 0.97 K ions per tetramer in G-quadruplexes prepared in KCl 0.5 M, while in all cases 6 ions per unit cell were detected to occupy the outer sites, partially neutralizing the two formal negative charges per phosphate group. The very large K ions concentration difference between the binding sites and the bulk solution demonstrates that counterions are structurally involved in the formation and in the stabilization of the helices.