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A hierarchical computational approach is used to identify the engineered binding-site cavity at the remodeled intermolecular interface between the mutants of human growth hormone (hGH) and the extracellular domain of its receptor (hGHbp). Multiple docking simulations are conducted with the remodeled hGH-hGHbp complex for a panel of potent benzimidazole-containing inhibitors that can restore the binding affinity of the wild-type complex, and for a set of known nonactive small molecules that contain different heterocyclic motifs. Structural clustering of ligand-bound conformations and binding free-energy calculations, using the AMBER force field and a continuum solvation model, can rapidly locate and screen numerous ligand-binding modes on the protein surface and detect the binding-site hot spot at the intermolecular interface. Structural orientation of the benzimidazole motif in the binding-site cavity closely mimics the position of the hot spot residue W104 in the crystal structure of the wild-type complex, which is recognized as an important structural requirement for restoring binding affinity. Despite numerous pockets on the protein surface of the mutant hGH-hGHbp complex, the binding-site cavity presents the energetically favorable hot spot for the benzimidazole-containing inhibitors, whereas for a set of nonactive molecules, the lowest energy ligand conformations do not necessarily bind in the engineered cavity. The results reveal a dominant role of the intermolecular van der Waals interactions in providing favorable ligand-protein energetics in the redesigned interface, in agreement with the experimental and computational alanine scanning of the hGH-hGHbp complex., (Copyright 2003 Wiley-Liss, Inc.)