Many biomedical applications, such as classification of binding specificities or bioengineering, depend on the accurate definition of protein binding interfaces. Depending on the choice of method used, substantially different sets of residues can be classified as belonging to the interface of a protein. A typical approach used to verify these definitions is to mutate residues and measure the impact of these changes on binding. Besides the lack of exhaustive data, this approach also suffers from the fundamental problem that a mutation introduces an unknown amount of alteration into an interface, which potentially alters the binding characteristics of the interface. In this study we explore the impact of alternative binding site definitions on the ability of a protein to recognize its cognate ligand using a pharmacophore approach, which does not affect the interface. The study also shows that methods for protein binding interface predictions should perform above approximately F-score = 0.7 accuracy level to capture the biological function of a protein.