Interaction strength of osmolytes with the anion of a salt-bridge determines its stability

In order to understand the role of osmolytes in regulating physicochemical behavior of proteins, we investigated the influence of protein destabilizing (urea and guanidinium chloride) and stabilizing osmolytes (TMAO, glycerol, and betaine) on a model salt-bridge (SB) formed between structural analogues of arginine and glutamate/aspartate sidechains in a solvent continuum using first-principles quantum chemical calculations based on DFT and MP2 methods. The binding strength of the osmolyte with the SB is found to be in the order of betaine > TMAO > Gdm+ > glycerol > urea. The osmolytes (TMAO and betaine) that preferentially bind to the SB cation have a marginal influence on SB stability. Also, pure π–π stacking interaction between Gdm+ and the SB cation plays an insignificant role in destabilizing the SB. In fact, the interaction strength of osmolytes with the SB anion mainly determines the stability of SB. For instance, a competition between Gdm+ and the SB cation to bind with the SB anion is responsible for instability and subsequent dissociation of the SB. The competition provided by other osmolytes is too weak to break the SB. Exploiting this information, we designed three structural derivatives of Gdm+, all having a stronger interaction with SB anion, and thereby show a stronger SB dissociation potential. Furthermore, we find an excellent linear anti-correlation between SB interaction energy and the energy of interaction between osmolyte and the SB anion, which suggests that by knowing only the strength of osmolyte⋯acetate interaction, one can predict the influence of osmolytes on the salt-bridge instability. This information is useful in fine-tuning the SB dissociation power of Gdm+, which has a practical significance in obtaining the mechanistic insight into the influence of GdmCl on protein stability. Our results also provide a basis for understanding the chemistry of other ion-pairs formed between a cationic hydrogen donor and an anionic acceptor.