Molecular crowding effect on specific binding of Hg²⁺ to T-T mismatched base pair in duplex DNA

Many biomolecules are crowded in vivo environments. Metal ion-nucleic acid interactions are important in vivo molecular crowding conditions for structure formation and biological activity of nucleic acids. Although metal ion-nucleic acid interactions have been investigated in detail under diluted conditions, studies examining the molecular crowding effect on metal ion-nucleic acid interactions are limited. Hg²⁺ specifically binds to T-T mismatched duplex DNA to form T-Hg-T base pair under diluted conditions. Here, we examined the binding under molecular crowding conditions. To the best of our knowledge, no previous studies reported the metal-mediated base-pair formation under molecular crowding conditions. UV melting showed that the specific stabilization of only the T-T mismatched duplex by Hg²⁺ addition was maintained under molecular crowding conditions. CD spectra showed that no significant structural change of the T-T mismatched duplex by Hg²⁺ addition was preserved under molecular crowding conditions. Isothermal titration calorimetric analyses showed that the 1:1 M ratio for the specific binding of Hg²⁺ to T-T was maintained under molecular crowding conditions. However, the magnitudes of the negative ∆H and the positive ∆S were significantly larger and smaller, respectively, than those under diluted conditions, which may lead to the smaller magnitudes of K_a and ∆G. Smaller number of released water molecules upon the binding under molecular crowding conditions may result in these results. The present findings may be useful for developing efficient metal-mediated base-pair formation, leading to progress in their efficient applications in various fields including nanotechnology.