Nilapwar, S.; (2009) Characterization and exploitation of protein ligand interactions for structure based drug design. Doctoral thesis , UCL (University College London).

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Abstract

Most characterised protein-small molecule interactions that display a change in heat capacity (\bigtriangleupCp) occur with a negative \bigtriangleupCp value. This is often attributed to solvent reorganisation from reduction in solvent accessible apolar surface area accompanying complex formation. Positive \bigtriangleupCp values have not been widely reported and could typically be attributed to an increased solvent accessible apolar surface area, desolvation of polar surface area or structural transitions in the biomolecular complex. Heat shock protein-90 (Hsp90) is one of the abundant and important molecular ATP-dependent chaperones. The N-terminal domain of Hsp90 contains ATP/ADP binding site, where Hsp90-ADP interactions proceed with a large positive \bigtriangleupCp of 2.35 ± 0.46 kJ·mol-1·K-1. Interestingly geldanamycin, an Hsp90 inhibitor which binds to the same N-Hsp90-ADP/ATP binding site, interacts with a negative \bigtriangleupCp of -0.39 ± 0.04 kJ·mol-1·K-1. The semi-empirical correlation of the solvent accessible surface area change does not match well with the observed \bigtriangleupCp. This prompted us to investigate various factors affecting the thermodynamics of protein-small molecule binding including varying buffers, differing salt concentration, altering pH, substitution of different metal cations and performing interactions in heavy water. Molecular dynamics simulation and NMR studies have allowed us to disregard structural changes of N-Hsp90-ADP molecule from giving rise to positive \bigtriangleupCp. From a combination of these calorimetric, simulation and structural studies we have gathered a considerable body of evidence suggesting that the change in accessible surface area, ionic interactions and resultant desolvation of water molecules from the surface of a Mg2+ ion can contribute substantially to a positive \bigtriangleupCp. We conclude that this unique result appears to come from extensive disruption of the tightly bound water molecules present around Mg2+-ADP after binding to Hsp90, which then gives rise to a positive \bigtriangleupCp. In addition to these findings, the thermodynamics of 18 structurally related CDK2 inhibitors were investigated using ITC. CDK2 is a member of cyclin dependent kinases implicated in eukaryotic cell cycle progression and control. This investigation showed that even conservative changes in small molecule structure can reveal large variation in thermodynamic signature, while simple concepts such as van der Waals interactions, steric hindrance, and hydrophobicity are insufficient to explain it.

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