ИСТИНА

Structural stability and other aspects of physical chemistry of aptamers are important issues for the pharmacological industry and modern biology. The formation and structural stability of aptamers depend on several factors, such as stacking interactions between nucleic acid bases, hydrogen bonds between them, electrostatic interactions and hydration shell. In addition, some aptamers containing quadruplexes have also specific stability factors, namely, coordination of carbonyl oxygens by cations inside the G-DNA stems, and presence of connecting single-stranded loops in monomeric and dimeric G-DNA molecules. Loops' length and sequence have a strong influence on G-quadruplex stability and folding efficiency. One of primary tasks in development of therapeutic agent on base of aptamer is to optimize and unify the folding process. For example quadruplex substructure depends on cations size and charge. Most of small nucleic acid structures have been solved with NMR which has limited capability to detect stabilizing ions. In the present study on thrombin aptamer (15-TBA), a combination of explicit solvent molecular dynamics simulation (30 simulations, 4 µs in total), hybrid quantum mechanics/molecular mechanics approach and isothermal titration calorimetry was used to investigate the atomistic picture of ion binding to 15-TBA. We found that the individual ion binding events are effected by connecting loops, which play several roles. They stabilize the molecule during time periods when the bound ions are absent, they modulate the route of the ion into the G-stem, and they also stabilize, already coordinated ions by closing the gates through which the ions enter the quadruplex. Our observations leads to obvious modifications in aptamer structure. Newly synthesized seqences was tested and showed better anticoagulant activity and folding efficiency.