In the context of drug design, C-H···O hydrogen bonds have received little attention so far, mostly because they are considered weak relative to other noncovalent interactions such as O-H···O hydrogen bonds, π/π interactions, and van der Waals interactions. Herein, we demonstrate the significance of hydrogen bonds between C-H groups adjacent to an ammonium cation and an oxygen atom (N+-C-H···O hydrogen bonds) in protein-ligand complexes. Quantum chemical calculations revealed details on the strength and geometrical requirements of these N+-C-H···O hydrogen bonds, and a subsequent survey of the Protein Data Bank (PDB) based on these criteria suggested that numerous protein-ligand complexes contain such N+-C-H···O hydrogen bonds. An ensuing experimental investigation into the G9a-like protein (GLP)-inhibitor complex demonstrated that N+-C-H···O hydrogen bonds affect the activity of the inhibitors against the target enzyme. These results should provide the basis for the use of N+-C-H···O hydrogen bonds in drug discovery.

We performed protein-ligand docking simulations with a ligand T247, which has been reported as a selective inhibitor of a histone deacetylase HDAC3, by the replica-exchange umbrella sampling method in order to estimate the free energy profiles along ligand docking pathways of HDAC3-T247 and HDAC2-T247 systems. The simulation results showed that the docked state of the HDAC3-T247 system is more stable than that of the HDAC2-T247 system although the amino-acid sequences and structures of HDAC3 and HDAC2 are very similar. By comparing structures obtained from the simulations of both systems, we found the difference between structures of hydrophobic residues at the entrance of the catalytic site. Moreover, we performed conventional molecular dynamics simulations of HDAC3 and HDAC2 systems without T247, and the results also showed the same difference of the hydrophobic structures. Therefore, we consider that this hydrophobic structure contributes to the stabilization of the docked state of the HDAC3-T247 system. Furthermore, we show that Tyr209, which is one of the hydrophobic residues in HDAC2, plays a key role in the instability from the simulation results of a mutated-HDAC2 system.

dichroism spectroscopy (CD) and nuclear magnetic resonance (NMR) spectroscopy. The ITC data reveal that N-MdmX2C exhibits relatively-strong affinity binding to nutlin-3a compared to N-MdmX. The CD analysis indicates that both N-MdmX2C and N-MdmX have similar secondary structure content while their tertiary structures show difference. However, the 15N-1H HSQC NMR spectra reveal that the conformation of N-MdmX2C becomes more flexible compared to N-MdmX unexpectedly. Nevertheless, the 15N-1H HSQC NMR titration confirms that N-MdmX2C has higher binding affinity for nutlin-3a than N-MdmX. On the other hand, both N-MdmX2C and N-MdmX exhibit the same binding affinity to p53p investigated with ITC and NMR titration. Taken together, the current results suggest the confirmation of N-MdmX2C resembles a binding intermediate state for ligand binding, providing an ideal model for anticancer drug design targeting MdmX/Mdm2.