title: Boosting Affinity by Correct Ligand Preorganization for the S2 Pocket of Thrombin: A Study by Isothermal Titration Calorimetry, Molecular Dynamics, and High-Resolution Crystal Structures.
abstract: Structural preorganization to fix bioactive conformations at protein binding sites is a popular strategy to enhance binding affinity during late-stage optimization. The rationale for this enhancement relates to entropic advantages assigned to rigidified versus flexible ligands. We analyzed a narrow series of peptidomimetics binding to thrombin. The individual ligands exhibit at P2 a conformationally flexible glycine, more restricted alanine, N-methylglycine, N-methylhomoalanine, and largely rigidified proline moiety. Overall, affinity was found to increase by a factor of 1000, explained partly by an entropic advantage. All ligands adopt the same binding mode with small deviations. The residual mobility of the bound ligands is decreased across the series, and a protein side chain differs in its order/disorder behavior along with changes in the surface-water network pattern established across the newly generated protein-ligand surfaces. The enthalpy/entropy inventory displays a rather complex picture and emphasizes that thermodynamics can only be compared in terms of relative differences within a structurally similar ligand series.
authors.affiliation: Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6, 35037, Marburg, Germany.
authors.forename: Eggert H
authors.initials: EH
authors.lastname: Rühmann
authors.affiliation: Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6, 35037, Marburg, Germany.
authors.forename: Melinda
authors.initials: M
authors.lastname: Rupp
authors.affiliation: Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6, 35037, Marburg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Betz
authors.affiliation: Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6, 35037, Marburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Heine
authors.affiliation: Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6, 35037, Marburg, Germany. klebe@mailer.uni-marburg.de.
authors.forename: Gerhard
authors.identifier: http://orcid.org/0000-0002-4913-390X
authors.initials: G
authors.lastname: Klebe
mesh_terms: D002151:Calorimetry; D018360:Crystallography, X-Ray; D008024:Ligands; D056004:Molecular Dynamics Simulation; D015394:Molecular Structure; D013816:Thermodynamics; D013917:Thrombin / Q000737:chemistry*
chemical_list: D008024:Ligands; D013917:Thrombin
keywords: crystal structure analysis; enthalpic binding signature; entropic binding signature; ligand optimization; preorganization; thermodynamic binding profile; thrombin