Thermodynamic Consequences of Tyr to Trp Mutations in the Cation-π-Mediated Binding of Trimethyllysine by the HP1 Chromodomain

Evolution has converged on cation-π interactions for recognition of quaternary alkyl ammonium groups such as trimethyllysine (Kme3). While computational modelling indicates that Trp provides the strongest cation-π interaction of the native aromatic amino acids, there is limited corroborative data from measurements within proteins. Herein we investigate a Tyr to Trp mutation in the binding pocket of HP1 chromodomain, a reader protein that recognizes Kme3. Binding studies demonstrate that the Trp-mediated cation-π interaction is about -5 kcal/mol stronger, while the Y24W crystal structure shows that the mutation is not perturbing. Quantum mechanical calculations indicate that greater enthalpic binding is predominantly due to increased cation-π interactions. NMR studies indicate that differences in the unbound state of the Y24W mutation lead to enthalphy-entropy compensation. These results provide direct experimental quantification of Trp versus Tyr in a cation-π interaction and afford insight into the conservation of aromatic cage residues in Kme3 reader domains.